JP2018501228A - Pirin polypeptide and immune modulation - Google Patents

Abstract

The present invention relates to a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression comprising said polynucleotide sequence for use in the treatment and / or prevention of a disorder of interest. A host cell comprising a vector, wherein the disorder is an inflammatory disorder and / or an autoimmune disorder; the polypeptide is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof A polypeptide wherein the polynucleotide sequence has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof And / or said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: Or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative thereof; about the polypeptide HP said polynucleotide sequence or said host cell.

Description

  The present invention relates to a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence for various therapeutic and nutritional uses A host cell comprising

  Bacteroides thetaiotaomicron has potent anti-inflammatory effects in vitro and in vivo (Kelly D, Campbell JI, King TP, Grant G, Jansson EA, Coutts AG, Pettersson S, Conway S. Commensal anaerobic) Nat Immunol. 2004 Jan; 5 (1): 104-12). gut bacteria attenuated inflammation by regulating nuclear-cytoplasmic shuttling of PPAR-gamma and RelA. Bacteroides tetaiotaomicron modulates the molecular signaling pathway of NF-κB (Kelly D, Campbell JI, King TP, Grant G, Jansson EA, Coutts AG, Pettersson S, Conway S. Commensal anaerobic gut bacteria attenuate inflammation by regulating nuclear-cytoplasmic shuttling of PPAR-gamma and RelA. Nat Immunol. 2004 Jan; 5 (1): 104-12). Specifically, Bacteroides tetaiotaomicron prevents activation of pro-inflammatory pathways by stopping the binding of the active component of NF-κB (RelA) to key genes in the nucleus (Kelly D, Campbell JI, King TP, Grant G, Jansson EA, Coutts AG, Pettersson S, Conway S. Commensal anaerobic gut bacteria attenuate inflammation by regulating nuclear-cytoplasmic shuttling of PPAR-gamma and RelA. Nat Immunol. 2004 Jan; 5 (1) : 104-12).

  B. The entire genome of B. thetaiotaomicron was sequenced and annotated in 2003 by the Gordon group (Washington University School of Medicine, USA) (Xu J, Bjursell MK, Himrod J, Deng S, Carmichael LK, Chiang HC, Hooper LV, Gordon JI. A genomic view of the human-Bacteroides thetaiotaomicron symbiosis. Science. 2003 Mar 28; 299 (5615): 2074-6).

Kelly D, Campbell JI, King TP, Grant G, Jansson EA, Coutts AG, Pettersson S, Conway S. Commensal anaerobic gut bacteria attenuate inflammation by regulating nuclear-cytoplasmic shuttling of PPAR-gamma and RelA. Nat Immunol. 2004 Jan; 5 (1): 104-12 Xu J, Bjursell MK, Himrod J, Deng S, Carmichael LK, Chiang HC, Hooper LV, Gordon JI.A genomic view of the human-Bacteroides thetaiotaomicron symbiosis. Science. 2003 Mar 28; 299 (5615): 2074-6

  Surprisingly, we are a virtual protein (HP, hypothetical protein; gene ID 1075517; gene name BT — 0187; accession number AA075294) identified from the genome of Bacteroides tetaiotaomicron (VPI5482), AAO752944.1. Has been found to reduce the inflammation of the cells.

  The present invention relates to a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression comprising said polynucleotide sequence for use in the treatment and / or prevention of a disorder of interest. A host cell comprising a vector, wherein the disorder is an inflammatory disorder and / or an autoimmune disorder, and the polypeptide is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof The polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof And / or the polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 Ku has at least 75% identity SEQ ID NO: 5 or a variant thereof, homologs, and fragments or derivatives thereof, said polypeptide HP, provides polynucleotide sequences or said host cell encoding said polypeptide HP.

  In another aspect, the present invention relates to a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, said polynucleotide sequence for use in modulating inflammation of a cell, tissue or organ of interest. Or a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide is at least 75 with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof. The polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof; and / Or the polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative thereof, wherein the polypeptide HP, provides polynucleotide sequences or said host cell encoding said polypeptide HP.

  In a further aspect, the invention provides a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, for use in improving the intestinal barrier integrity of a subject, Or a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof. And the polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or the polynucleotide The sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 Its variants, homologs having at least 75% identity with a fragment or derivative thereof, wherein the polypeptide HP, providing the polynucleotide sequence or the host cell.

  In another aspect, the present invention provides a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, for use in modifying a bacterial composition in a tissue or organ to provide a more beneficial microflora, A host cell comprising a polynucleotide sequence or a host cell comprising an expression vector comprising the polynucleotide sequence is provided. For example, the invention provides that the polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, wherein the polynucleotide sequence is SEQ ID NO: 2, which encodes a polypeptide having at least 75% identity with SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or One or more types of lactose-fermenting bacteria (eg, E. coli) in the tissue or organ of interest if they have at least 75% identity to SEQ ID NO: 5 or variants, homologs, fragments or derivatives thereof )) And / or one in the tissue or organ of interest. Or it may be useful to reduce the level of two or more types of non-lactose fermenting bacteria.

  In a further aspect, the present invention provides, in a further aspect, a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, for use in maintaining the length of the large and / or small intestine of a subject (eg, the subject has IBD), A host cell comprising a polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof The polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof And / or the polynucleotide sequence is SEQ ID NO: 1, sequence Issue 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative thereof, wherein the polypeptide HP, providing the polynucleotide sequence or the host cell.

  In a further aspect, the present invention encodes polypeptide HP, polypeptide HP for use in reducing disruption of the intestine (eg, large intestine) of a subject (eg, subject has IBD) A polynucleotide sequence, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof, Having at least 75% identity with a homologue, fragment or derivative, wherein said polynucleotide sequence has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof And / or the polynucleotide sequence is The polypeptide HP, the polynucleotide sequence or the host cell having at least 75% identity with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof.

  In another aspect, the present invention is used to modulate the expression of one or more pro-inflammatory genes and / or one or more barrier integrity genes in one or more cells of a subject. A polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence, wherein the polypeptide comprises the sequence SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof, homologs, fragments or derivatives having at least 75% identity, wherein said polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof A polypeptide having at least 75% identity with a homologue, fragment or derivative And / or wherein said polynucleotide sequence has at least 75% identity with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof A nucleotide sequence or said host cell is provided.

  The present invention, in another aspect, regenerating islet-derived 3 beta gene (Reg3b), resistin-like gamma-resistin-like beta in one or more cells of a subject (eg, subject has IBD) Gene (Retnlg | Retnlb), sucrase-isomaltase (alpha-glucosidase) gene (Si), defensin alpha 24 gene (Defa24), hydroxysteroid 11-beta dehydrogenase 2 gene (Hsd11b2), hydroxysteroid (17-beta) dehydrogenase 2 One or more selected from the group consisting of the gene (Hsd17b2), resistin-like molecule-beta (RELMb) and nuclear receptor 1D1 thyroid hormone receptor alpha gene (Nr1d1 | Thra) A host cell comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence for use in regulating the expression of a gene Wherein the polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, wherein the polynucleotide sequence is SEQ ID NO: 2, Encodes a polypeptide having at least 75% identity with SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or its variants, homologues, fragments or derivatives and few Provide said polypeptide HP, said polynucleotide sequence or said host cell having at least 75% identity.

  In a further aspect, the present invention provides a polypeptide HP, a polynucleotide sequence encoding a polypeptide HP, for use in reducing activation of a pro-inflammatory pathway in one or more cells of a subject, said polynucleotide A host cell comprising a sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide comprises at least SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof Encoding a polypeptide having 75% identity and wherein the polynucleotide sequence is at least 75% identical to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; And / or the polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative thereof, wherein the polypeptide HP, providing the polynucleotide sequence or the host cell.

  The invention in a further aspect, for use in reducing the activity and / or expression of NF-κβ in one or more cells (eg, epithelial cells, epidermal cells, neural cells and / or pancreatic cells) of a subject. A polypeptide cell, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence, wherein the polypeptide is SEQ ID NO: 2. SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof having at least 75% identity, wherein said polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues thereof A polypeptide having at least 75% identity with the fragment or derivative. And / or wherein said polynucleotide sequence has at least 75% identity with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof A nucleotide sequence or said host cell is provided.

  In another aspect, the present invention provides a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell for use in improving the health of a subject's gastrointestinal tract, A host cell comprising an expression vector comprising a polynucleotide sequence, wherein said polypeptide has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof The polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or the polynucleotide sequence SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or varian thereof , Homologs having at least 75% identity with a fragment or derivative thereof, wherein the polypeptide HP, providing the polynucleotide sequence or the host cell.

  In a further aspect, the present invention relates to a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence, and a pharmaceutically acceptable Wherein the polypeptide comprises at least 75% of SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof. And the polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or Or the polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: Or a variant thereof, homologs having at least 75% identity with a fragment or derivative, to provide the pharmaceutical composition.

  In a further aspect, the present invention relates to a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, and nutritionally permissive Wherein the polypeptide comprises at least 75% of SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof. And the polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or Or the polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: Or a variant thereof, homologs having at least 75% identity with a fragment or derivative, to provide the nutritional supplement.

  In another aspect, the invention provides a livestock feed comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence. A food, a dietary supplement or a food additive, wherein the polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof; The polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or the polynucleotide sequence comprises: SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a varian thereof , Homologs having at least 75% identity with a fragment or derivative thereof, wherein the animal feed, the food product, provides the nutraceutical or the food additive.

  The invention, in a further aspect, is a method for producing a pharmaceutical composition according to the invention, comprising a polypeptide HP, a polynucleotide sequence encoding a polypeptide HP, a host cell comprising said polynucleotide sequence, or said polynucleotide sequence Mixing a host cell containing an expression vector comprising a pharmaceutically acceptable excipient, carrier or diluent, wherein the polypeptide, polynucleotide sequence or host cell is encapsulated in the method. The polypeptide has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and the polynucleotide sequence is SEQ ID NO: 2, No. 4 or SEQ ID No. 6 or variants, homologues, fragments or derivatives thereof Encodes a polypeptide having at least 75% identity and / or the polynucleotide sequence is at least 75% identical to SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof There is provided a method as described above.

  In a further aspect, the present invention provides a method for producing a nutritional supplement according to the present invention, comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or said polynucleotide sequence. Mixing a host cell containing an expression vector comprising a nutritionally acceptable excipient, carrier or diluent, wherein the polypeptide or polynucleotide may be encapsulated by the method, The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, wherein said polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or sequence Number 6 or a variant, homolog, fragment or derivative thereof and at least 75 And / or the polynucleotide sequence has at least 75% identity with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof, The method is provided.

  In another aspect, the present invention provides a method for producing livestock feed, food, dietary supplement or food additive according to the present invention, comprising polypeptide HP, polynucleotide sequence encoding polypeptide HP, said polynucleotide sequence Or a host cell containing an expression vector containing the polynucleotide sequence is mixed with livestock feed, food, nutritional supplements, food additives or their ingredients, wherein the polypeptide or polynucleotide comprises Optionally encapsulated by said method, wherein said polypeptide has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, The sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof; Which encodes a polypeptide having at least 75% identity with a molog, fragment or derivative, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof The method is provided having at least 75% identity.

  In another aspect, the present invention provides a method for treating and / or preventing a disorder in a subject, comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or the poly Administering to said subject a host cell comprising an expression vector comprising a nucleotide sequence, said polypeptide or polynucleotide sequence or host cell treating and / or preventing said subject's disorder, said disorder being an inflammatory disorder And / or an autoimmune disorder, wherein the polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, wherein the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof, homologs, Or encoding a polypeptide having at least 75% identity with a derivative, and / or wherein said polynucleotide sequence is at least 75% with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof The method is provided having the same identity.

  The present invention, in a further aspect, is a method for modulating inflammation of a tissue or organ of interest comprising the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or said poly Administering to said subject a host cell comprising an expression vector comprising a nucleotide sequence, wherein said polypeptide, polynucleotide sequence or host cell modulates inflammation of said tissue or organ of said subject, said polypeptide comprising a sequence SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof, homologs, fragments or derivatives having at least 75% identity, wherein said polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof , Homologues, fragments or derivatives and at least 75% Encoding polypeptides having identity and / or wherein said polynucleotide sequence has at least 75% identity with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof, Provide a method.

  In another aspect, the present invention provides a method for improving intestinal barrier integrity of a subject, comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or said polynucleotide Administering to said subject a host cell comprising an expression vector comprising a sequence, wherein said polypeptide or polynucleotide sequence or host cell improves intestinal barrier integrity of said subject, said polypeptide comprising SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof having at least 75% identity, wherein said polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs thereof, Polype having at least 75% identity with the fragment or derivative Encoding the plastid, and / or the polynucleotide sequence, SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a variant thereof, having homologue, fragment or derivative thereof with at least 75% identity, to provide the method.

  The present invention, in another aspect, reduces the level of one or more types of lactose fermenting bacteria (eg, E. coli) in a subject tissue or organ and / or one or more in a subject tissue or organ. A method for reducing the level of two or more types of non-lactose fermenting bacteria comprising: polypeptide HP, polynucleotide sequence encoding polypeptide HP, host cell comprising said polynucleotide sequence, or said polynucleotide sequence Administering to said subject a host cell comprising said expression vector, wherein said polypeptide, polynucleotide sequence or host cell is one or more types of lactose-fermenting bacteria in said tissue or organ of interest ( For example, the level of E. coli) and / or the subject tissue or organ Reducing the level of one or more types of non-lactose fermenting bacteria in which the polypeptide is at least 75% of SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof And the polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or Or providing said method wherein said polynucleotide sequence has at least 75% identity to SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof.

  The present invention, in a further aspect, is a method for maintaining the length of the large and / or small intestine of a subject, comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or Administering to said subject (eg, the subject has IBD) a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide, polynucleotide sequence or host cell comprises said subject's large intestine and / or small intestine. The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, wherein the polynucleotide sequence has the sequence SEQ ID NO: 4, SEQ ID NO: 4 or SEQ ID NO: 6, or variants, homologues, fragments Encodes a polypeptide having at least 75% identity with the derivative, and / or the polynucleotide sequence is at least 75% with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof The method is provided having the same identity.

  In a further aspect, the present invention provides a method for reducing disturbances in the intestine (eg, large intestine) of a subject, the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or Administering to the subject (eg, the subject has IBD) a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide, polynucleotide sequence or host cell is intestinal (eg, large intestine) of said subject. The polypeptide has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, wherein the polynucleotide sequence has the sequence No. 2, SEQ ID No. 4 or SEQ ID No. 6 or variants, homologues, fragments thereof Or a polypeptide having at least 75% identity with a derivative and / or wherein said polynucleotide sequence is at least 75 with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof. The method is provided having% identity.

  In another aspect, the present invention provides a method for modulating the expression of one or more pro-inflammatory or anti-inflammatory genes in one or more cells of a subject, comprising the steps of polypeptide HP, polypeptide HP Administering to said subject a coding polynucleotide sequence, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide, polynucleotide sequence or host cell comprises: Modulates the expression of one or more pro- and / or anti-inflammatory genes in one or more cells of the subject, wherein the polypeptide comprises SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or The polynucleotide sequence having at least 75% identity with the variant, homologue, fragment or derivative thereof, Encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, sequence Said method is provided having at least 75% identity with No. 3 or SEQ ID No. 5 or variants, homologues, fragments or derivatives thereof.

  In another aspect, the invention provides a method of modulating the expression of one or more genes in one or more cells of a subject, comprising the polypeptide HP, or a polynucleotide sequence encoding the polypeptide HP. Administering to said subject a host cell that is administered to said subject or comprising a host cell comprising said polynucleotide sequence or comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide, polynucleotide sequence or host cell comprises Regulates the expression of one or more genes in one or more cells of the subject, wherein the one or more genes are regenerated islet-derived 3beta gene (Reg3b), resistin-like gamma-resistin-like Beta gene (Retnlg | Retnlb), sucrase-isomaltase (alpha- (Lucosidase) gene (Si), defensin alpha 24 gene (Defa24), hydroxysteroid 11-beta dehydrogenase 2 gene (Hsd11b2), hydroxysteroid (17-beta) dehydrogenase 2 gene (Hsd17b2), resistin-like molecule-beta (RELMb) and A nuclear receptor 1D1 thyroid hormone receptor alpha gene (Nr1d1 | Thra) selected from the group (eg, the subject has IBD) and the polypeptide is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or Having at least 75% identity with its variant, homologue, fragment or derivative, wherein said polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variant, homologue, fragment or derivative thereof And / or the polynucleotide sequence is at least 75% identical to SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof There is provided a method as described above.

  In a further aspect, the present invention provides a method for reducing activation of a pro-inflammatory pathway in one or more cells of a subject, comprising polypeptide HP, a polynucleotide sequence encoding polypeptide HP, said polynucleotide sequence Administering to the subject a host cell comprising or a host cell comprising an expression vector comprising the polynucleotide sequence, wherein the polypeptide, polynucleotide sequence or host cell induces inflammation in one or more cells of the subject Reducing the activation of the sexual pathway, wherein the polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, wherein the polynucleotide sequence SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof, Which encodes a polypeptide having at least 75% identity with a molog, fragment or derivative, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof The method is provided having at least 75% identity.

  In a further aspect, the present invention is a method of reducing the activity and / or expression of NF-κβ in one or more cells (eg, epithelial cells, epidermal cells, neural cells and / or pancreatic cells) of a subject, Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, A polynucleotide or host cell that reduces the activity and / or expression of NF-κβ in one or more cells of the subject (eg, epithelial cells, epidermal cells, neuronal cells and / or pancreatic cells); SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or inducers thereof A polypeptide having at least 75% identity to the body and wherein the polynucleotide sequence is at least 75% identical to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof The method is provided wherein it encodes and / or the polynucleotide sequence has at least 75% identity with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof.

  In another aspect, the present invention provides a method for improving the health of a subject's gastrointestinal tract, comprising polypeptide HP, a polynucleotide sequence encoding polypeptide HP, a host cell comprising said polynucleotide sequence, or said polynucleotide Administering to said subject a host cell comprising an expression vector comprising a sequence, wherein said polypeptide, polynucleotide sequence or host cell improves the gastrointestinal tract health of said subject, said polypeptide comprising SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof having at least 75% identity, wherein said polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs thereof, A polypeptide having at least 75% identity with a fragment or derivative Encoding, and / or the polynucleotide sequence, SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative, to provide the method.

  In a further aspect, the present invention provides, in a further aspect, a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, for the manufacture of a medicament for the treatment and / or prevention of a disorder of interest. Use of a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said disorder is an inflammatory disorder and / or an autoimmune disorder and said polypeptide is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 Or at least 75% identity with variants, homologues, fragments or derivatives thereof, wherein said polynucleotide sequence is at least 75% with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof A polypeptide having the identity of and / or said polynucleotide De sequence, SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative, to provide the use.

  In another aspect, the present invention provides a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, for producing a drug that modulates inflammation of a tissue or organ of interest. Or use of a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide is at least 75% identical to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof Wherein the polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or The polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative, to provide the use.

  In another aspect, the invention provides a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host for producing a drug that improves the intestinal barrier integrity of a subject, Use of a host cell comprising an expression vector comprising a polynucleotide sequence, wherein the polypeptide has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof. And the polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or the polynucleotide The sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Variants, homologs having at least 75% identity with a fragment or derivative, to provide the use.

  In a further aspect, the present invention provides, in a further aspect, a polypeptide HP, a polynucleotide sequence encoding a polypeptide HP, for producing a drug that maintains the length of the large and / or small intestine of a subject (eg, the subject has IBD) , A host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence, wherein the polypeptide comprises SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof, homologs The polynucleotide sequence has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof. Encoding a polypeptide and / or wherein said polynucleotide sequence is No. 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative, to provide the use.

  In a further aspect, the present invention provides a drug that modifies the bacterial composition in a tissue or organ to provide a beneficial microbiota, preferably one or more types of lactose-fermenting bacteria in the tissue or organ of interest. To produce a drug for use in reducing the level of (eg, E. coli) and / or reducing the level of one or more types of non-lactose fermenting bacteria in a tissue or organ of interest. Of the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence, wherein the polypeptide comprises the sequence SEQ ID NO: 4, SEQ ID NO: 4 or SEQ ID NO: 6, or variants, homologues, fragments or derivatives thereof At least 75% identity and the polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof And / or provides said use wherein said polynucleotide sequence has at least 75% identity with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof.

  In another aspect, the present invention relates to a polypeptide HP, a polynucleotide encoding a polypeptide HP, for producing a drug that reduces disturbances in the intestine (eg, the large intestine) of a subject (eg, the subject has IBD). Use of a host cell comprising a sequence, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence, wherein said polypeptide comprises SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof, Having at least 75% identity with a homologue, fragment or derivative, wherein said polynucleotide sequence has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof And / or the polynucleotide sequence is a sequence Issue 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative, to provide the use.

  In a further aspect, the present invention provides a polypeptide HP, a polypeptide encoding a polypeptide HP, for producing a drug that modulates the expression of one or more pro-inflammatory genes in one or more cells of a subject. Use of a nucleotide sequence, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof Having at least 75% identity with a homologue, fragment or derivative, wherein said polynucleotide sequence is at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof And / or said polynucleotide sequence But SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative, to provide the use.

  The invention, in another aspect, regenerates islet-derived 3 beta gene (Reg3b), resistin-like gamma-resistin-like beta gene (Retnlg | Retnlb), sucrase in one or more cells of a subject (eg, subject has IBD) -Isomaltase (alpha-glucosidase) gene (Si), defensin alpha 24 gene (Defa24), hydroxysteroid 11-beta dehydrogenase 2 gene (Hsd11b2), hydroxysteroid (17-beta) dehydrogenase 2 gene (Hsd17b2), resistin-like molecule A drug that regulates the expression of one or more genes selected from the group consisting of beta (RELMb) and nuclear receptor 1D1 thyroid hormone receptor alpha gene (Nr1d1 | Thra) Use of a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence for the construction of said polypeptide Having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, wherein said polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 Or a polypeptide that has at least 75% identity with a variant, homolog, fragment or derivative thereof, and / or the polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologs thereof, At least 75% identical to the fragment or derivative The a, providing the use.

  In another aspect, the invention provides a polypeptide HP, a polynucleotide sequence encoding a polypeptide HP, for producing a drug that reduces activation of a pro-inflammatory pathway in one or more cells of a subject, Use of a host cell comprising a nucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, wherein said polypeptide is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or A polypeptide having at least 75% identity with a derivative, wherein the polynucleotide sequence has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof. And / or the polynucleotide sequence is SEQ ID NO: 1, No. 3 or SEQ ID NO: 5 or a variant thereof, homologs having at least 75% identity with a fragment or derivative, to provide the use.

  In a further aspect, the present invention provides a medicament for reducing the activity and / or expression of NF-κβ in one or more cells (eg, epithelial cells, epidermal cells, neural cells and / or pancreatic cells) of a subject. Of the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence, wherein the polypeptide comprises the sequence SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof, homologs, fragments or derivatives having at least 75% identity, wherein said polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof A polypeptide having at least 75% identity with a homologue, fragment or derivative The use is provided wherein it encodes a tide and / or the polynucleotide sequence has at least 75% identity to SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof.

  In another aspect, the present invention relates to a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell for producing a drug that improves the health of the subject's digestive tract. Use of a host cell comprising an expression vector comprising a polynucleotide sequence is provided.

The present invention will be described with reference to the accompanying drawings.
A polynucleotide sequence encoding HP (SEQ ID NO: 1), a polynucleotide sequence encoding E. coli optimized HP (Rec 1 HP-SEQ ID NO: 3) and L. FIG. 5 shows an alignment of polynucleotide sequence (Rec 2 HP—SEQ ID NO: 5) encoding L. lactis optimized HP. HP has been deposited with GenBank under the accession number AAO75294.1 and is described in GenBank as a putative Pirin family protein [Bacteroides tetaiotaomicron VPI-5482]. Polypeptide sequence HP (SEQ ID NO: 2), E. coli optimized HP (Rec 1 HP-SEQ ID NO: 4) and L. FIG. 4 shows an alignment of lactis optimized HP (Rec 2 HP—SEQ ID NO: 6). HP has been deposited with GenBank under the accession number AAO75294.1 and is described in GenBank with the putative Pirin family protein [Bacteroides tetaiotaomicron VPI-5482]. Changes in body weight, water and food intake by rats fed with dextran sodium sulphate (DSS) dissolved in water with (DSS / HP) or without (DSS) hypothetical protein (HP) FIG. FIG. 2 shows the length of the colon and small intestine in rats given dextran sulfate sodium dissolved in water with (DSS / HP) or not (DSS) combined with a hypothetical protein (HP). Lactose-fermenting bacteria (mainly E. coli) in tissues from rats given dextran sulfate sodium (DSS) dissolved in water with (DSS / HP) or without (DSS) combined with hypothetical protein (HP) and It is a figure which shows the number of non-lactose fermentative bacteria. [No bacteria were detected when Log 10 = 1.0]. FIG. 4 shows the morphology of the ascending and descending colons from rats fed with dextran sulfate sodium dissolved in water with (DSS / HP) or without (DSS) hypothetical protein (HP). Mean histopathology score for ascending colon from rats given dextran sulfate sodium dissolved in water with or without hypothetical protein (HP) (DSS / HP) or (DSS), and grades 0-3 FIG. 6 is a diagram showing a histopathological diagnosis score as a percentage of the visual field regarding the pathological condition. FIG. 4 shows a heat map of 377 genes that are differentially expressed in tissues from rats fed with sodium dextran sulfate dissolved in water, with or without hypothetical protein (HP). Diagram showing expression of inflammation-related genes (as real-time PCR) in ascending colon from rats given dextran sulfate sodium dissolved in water with or without virtual protein (HP) (DSS / HP) or without (DSS) It is.

HP
While not wishing to be bound by theory, the polypeptide HP of the present invention is a pilin related protein.

  The polypeptide HP is at least 75%, 80%, 85%, 90%, 95%, 97% of the polypeptide sequence shown as SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof. , 98%, 99% or 100% identity.

An example of a polypeptide HP of the invention is SEQ ID NO: 2 deposited at GenBank as AAO75294.1, and Bacteroides tetaiotaomicron containing SEQ ID NO: 1 has been deposited at DSM2079 [E50 (VPI5482), VPI5482]. Can be found in the German microbial cell culture collection limited company (DSMZ, Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH). The polypeptide sequence SEQ ID NO: 2 has the following sequence:
1 0 2 0 3 0 4 0 5 0 6 0
MKKVIDRASS RGYFNHGWLK THHTFSFANY YNPERIHFGA LRVLNDDSVD PSMGFDTHPH

7 0 8 0 9 0 10 0 11 0 12 0
KNMEVISIPL KGYLRHGDSV QNTKTITPGD IQVMSTGSGI YHSEYNDSKE EQLEFLQIWV

13 0 14 0 15 0 16 0 17 0 18 0
FPRIENTKPE YNNFDIRPLL KPNELSLFIS PNGKTPASIK QDAWFSMGDF DTERTIEYCM

19 0 20 0 21 0 22 0 23 0
HQEGNGAYLF VIEGEISVAD EHLAKRDGIG IWDTKSFSIR ATKGTKLLVM EVPM

  AAO75294.1 has been described as a putative Pirin family protein. AAO75294.1 was identified from Bacteroides tetaiotaomicron VPI-5482.

  The polypeptide sequences deposited with GenBank as AAO768683.1 and CDE805552.1 are examples of polypeptides having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6.

The polypeptide sequence of AAO768683.1 is as follows:
1 0 2 0 3 0 4 0 5 0 6 0
MKKVIHKADT RGHSQYDWLD SYHTFSFDEY FDSDRINFGA LRVLNDDKVA PGEGFQTHPH

7 0 8 0 9 0 10 0 11 0 12 0
KNMEIISIPL KGHLQHGDSK KNSRIITVGE IQTMSAGTGI FHSEVNASPV EPVEFLQIWI

13 0 14 0 15 0 16 0 17 0 18 0
MPRERNTHPV YKDFSIKELE RPNELAVIVS PDGSTPASLL QDTWFSIGKV EAGKKLGYHL

19 0 20 0 21 0 22 0 23 0
HQSHGGVYIF LIEGEIVVDG EVLKRRDGMG VYDTKSFELE TLKDSHILLI EVPM

  The polypeptide sequence of AAO768683.1 is also referred to herein as SEQ ID NO: 7. AAO768683.1 is described in GenBank as a putative Pirin family protein. AAO768683.1 was isolated from Bacteroides tetaiotaomicron VPI-5482. [E50 (VPI 5482), VPI 5482]. Bacteroides tetaiotaomicron containing SEQ ID NO: 7 can be found in DSMZ (German Microbial Cell Culture Collection), deposited as DSM 2079 BT 1576.

The polypeptide sequence of CDE805552.1 is as follows:
10 20 30 40 50 60
MKKVIHKADT RGHSQYDWLD SYHTFSFDEY FDSDRINFGA LRVLNDDKVA PGEGFQTHPH

70 80 90 100 110 120
KNMEIISIPL KGHLQHGDSK KNSRIITVGE IQTMSAGTGI FHSEVNASPV EPVEFLQIWI

130 140 150 160 170 180
MPRERNTHPV YKDFSIKELE RPNELAVIVS PDGSTPASLL QDTWFSIGKV EAGKKLGYHL

190 200 210 220 230
HQSHGGVYIF LIEGEIVVDG EVLKRRDGMG VYDTKSFELE TLKDSHILLI EVPM

  The polypeptide sequence of CDE805552.1 is also referred to herein as SEQ ID NO: 8. CDE805552.1 is described in GenBank as a putative Pirin family protein. CDE805552.1 was isolated from Bacteroides tetaiotaomicron CAG: 40.

  The terms “polypeptide HP”, “HP polypeptide” and “HP” are used synonymously herein.

  In one embodiment, the polypeptide HP is the polypeptide shown as SEQ ID NO: 2.

  In another embodiment, the polypeptide HP is the polypeptide shown as SEQ ID NO: 4.

  In a further embodiment, the polypeptide HP is the polypeptide set forth as SEQ ID NO: 6.

  HP polypeptides can be obtained from certain microorganisms. In one embodiment, the HP polypeptide is obtained from an anaerobic, gram-negative bacterium that can live in the gastrointestinal tract. In a further embodiment, the HP polypeptide is obtained from Bacteroides spp, such as Bacteroides tetaiotaomicron.

  Examples of polynucleotide sequences encoding the polypeptide HP include the polynucleotide sequence shown as SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5, and the polypeptide shown as SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6. And at least 75%, 80%, 85%, 90%, 95%, 97%, 98% or 99% with a polynucleotide sequence, SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologs, fragments or derivatives thereof At least 75%, 80%, 85%, 90%, 95% to a polypeptide sequence having the identity of SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or a variant, homolog, fragment or derivative thereof A polypeptide encoding a polypeptide having 97%, 98% or 99% identity Kureochido sequence, and polynucleotide sequences include encoding SEQ ID NO: 7 or SEQ ID NO: 8.

  SEQ ID NOs: 1, 3 and 5 are shown in FIG. 1A.

SEQ ID NO: 1 has the following sequence:
Atgaaaaaagtaatcgacagagcttcatcaagaggctattttaatcatggctggctcaaaacccaccacacattcagttttgctaactattacaatccggaaagaatccatttcggagccttgcgagtgctgaatgatgacagtgtagacccgtcgatgggatttgatactcatccacataaaaatatggaagtaatttccattccgttgaaagggtatctgagacatggcgacagtgtacaaaatacgaaaacgattactcccggtgatatccaagtgatgagtacgggcagtggtatctatcatagtgagtataacgacagcaaggaagaacaattggaattcctgcaaatatgggtattcccccgaatcgagaatacgaaacccgaatataacaatttcgatatacgtccgctgctgaaaccgaacgagttatctctgttcatttcaccgaacggcaagacaccggcctccatcaaacaggatgcctggttctctatgggagacttcgatacggaaagaaccatcgaatattgtatgcatcaggaaggtaacggagcttatctgtttgtgatagaaggagagatcagcgtggccgatgaacatctggccaaacgtgacggcatcggaatatgggataccaaaagcttctctatccgtgctactaaagggaccaaacttctggtaatggaagtacccatgtaa

  SEQ ID NO: 1 encodes SEQ ID NO: 2 deposited at GenBank with accession number AAO75294.1.

  The polynucleotide sequence of SEQ ID NO: 1 was codon optimized for expression in E. coli. This codon optimized sequence is shown as SEQ ID NO: 3. This sequence is sometimes referred to herein as “Rec 1 HP” or “Recombinant 1 HP”.

SEQ ID NO: 3 has the following sequence:
ggtaccatgaaaaaagtgattgatcgtgcgagcagccgtggctattttaaccatggctggctgaaaacccatcatacctttagcttcgcgaactattataatccggaacgcattcattttggcgcgctgcgtgtgctgaacgatgatagcgtggatccgagcatgggctttgatacccatccgcacaaaaacatggaagtgattagcattccgctgaaaggctatctgcgtcatggcgatagcgtgcagaacaccaaaaccattaccccgggtgatattcaggtgatgagcaccggcagcggcatttatcatagcgaatacaacgatagcaaagaagaacagctggaatttctgcagatttgggtgtttccgcgtattgaaaacaccaaaccggaatataacaactttgatattcgcccgctgctgaaaccgaacgaactgagcctgtttattagcccgaacggcaaaaccccggcgagcattaaacaggatgcgtggtttagcatgggcgattttgataccgaacgcaccattgaatattgcatgcatcaggaaggcaacggcgcgtacctgtttgtgattgaaggcgaaattagcgtggcggatgaacatctggccaaacgtgatggcattggcatttgggataccaaaagcttcagcattcgtgcgaccaaaggcaccaaactgctggtgatggaagtgccgatgtaataagagctc

The polypeptide sequence encoded by SEQ ID NO: 3 is shown as SEQ ID NO: 4. SEQ ID NO: 4 has the following sequence:
GTMKKVIDRASSRGYFNHGWLKTHHTFSFANYYNPERIHFGALRVLNDDSVDPSMGFDTHPHKNMEVISIPLKGYLRHGDSVQNTKTITPGDIQVMSTGSGIYHSEYNDSKEEQLEFLQIWVFPRIENTKPEYNNFDIRPLLKPNEGSVYLFLFNGKTPEGIK

  The polynucleotide sequence of SEQ ID NO: 1 was codon optimized for expression in Lactococcus lactis. This codon optimized sequence is shown as SEQ ID NO: 5. This sequence is sometimes referred to herein as “Rec 2 HP” or “Recombinant 2 HP”.

SEQ ID NO: 5 has the following sequence:
ggtaccatgaaaaaagttattgatcgtgcttcatcacgtggatattttaatcatggatggcttaaaactcatcatacatttagttttgccaattattataatccagaacgtattcattttggtgctcttcgtgttcttaatgatgattcagttgatccatcaatgggatttgatacacatccacataaaaatatggaagttatttcaattccacttaaaggatatcttcgtcatggtgattcagttcaaaatacaaaaacaattacacctggagatattcaagttatgtctacaggatcaggaatttatcattcagaatataatgattcaaaagaagaacaacttgaatttcttcaaatttgggtctttccacgtattgaaaatacaaaaccagaatataataatttcgacattcgtccacttcttaaaccaaatgaactttcactttttatctcaccaaatggaaaaacaccagcttcaattaaacaagatgcttggttttcaatgggagattttgatacagaacgtacaattgaatattgtatgcatcaagaaggtaacggcgcttatctttttgttattgaaggtgaaatttcagttgctgatgaacatcttgctaaacgtgatggaattggaatttgggatacaaaatcattttcaattcgtgctacaaaaggtacaaaacttcttgttatggaagttccaatgtaataagagctc

The polypeptide sequence encoded by SEQ ID NO: 5 is shown as SEQ ID NO: 6. SEQ ID NO: 6 has the following sequence:
GTMKKVIDRASSRGYFNHGWLKTHHTFSFANYYNPERIHFGALRVLNDDSVDPSMGFDTHPHKNMEVISIPLKGYLRHGDSVQNTKTITPGDIQVMSTGSGIYHSEYNDSKEEQLEFLQIWVFPRIENTKPEYNNFDIRPLLKPNEGSVYLFLFNGKTPEGIK

  In one embodiment, the polynucleotide sequence encoding the polypeptide HP is at least 75%, 80% of the polynucleotide sequence shown as SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof, 85%, 90%, 95%, 97%, 98% or 99% identity.

  In one embodiment, the polynucleotide sequence encoding the polypeptide HP is a polypeptide shown as SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6, or SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants thereof, homologs , Encodes a polypeptide having at least 75%, 80%, 85%, 90%, 95%, 97%, 98% or 99% identity to the polypeptide shown as a fragment or derivative.

  In one embodiment, the polypeptide HP is a truncated HP polypeptide. For example, the truncated polypeptide comprises at least 20, 30, 40, 50, 75, 100, 125, 150, 175 or 200 amino acids of the polypeptide set forth as SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6.

  In one embodiment, the polynucleotide sequence encoding polypeptide HP encodes a truncated HP polypeptide.

  In one embodiment, the polypeptide HP is a fusion polypeptide. For example, the polypeptide is fused to glutathione S-transferase (GST).

Host cell In one aspect, a host cell described herein comprises a polynucleotide sequence encoding the polypeptide HP.

  In another embodiment, the host cell described herein comprises an expression vector comprising a polynucleotide sequence that encodes the polypeptide HP.

  In further embodiments, a host cell described herein has been transformed with a nucleotide sequence that causes the host cell to overexpress HP. For example, a promoter is inserted into the genome of a host cell, thereby allowing the host cell to overexpress a polynucleotide sequence HP (eg, an endogenous polynucleotide sequence), ie, the promoter is a polynucleotide encoding HP. The sequence can be overexpressed.

  As used herein, the term “overexpressing” in the phrases “a nucleotide sequence that causes a host cell to overexpress HP” and “a promoter that can overexpress a polynucleotide sequence encoding HP” Refers to an increase in expression from zero to a certain expression level, or a transition from a lower expression level to a higher expression level (eg, upregulation), as compared to a comparable host cell prior to transformation. .

  In one embodiment, the level of mRNA encoding HP in a transformed host cell that overexpresses HP is such that the level of mRNA is at least 10% in the transformed host cell compared to an equivalent host cell prior to transformation, Increased by 20%, 30%, 40% or 50% (ie, up-regulated).

  Examples of host cells that overexpress HP include the following: (i) host cells transformed with an expression vector encoding HP (the host cells may express HP before transformation). And (ii) a host cell transformed to upregulate endogenous HP expression (before transformation, the host cell overexpresses the HP for a given set of culture conditions). But after transformation, the host cells can overexpress the HP at higher levels under the same culture conditions).

  The polynucleotide sequence encoding the polypeptide HP may be codon optimized for the host cell. For example, the polypeptide sequence may be codon optimized for expression in E. coli (eg, SEQ ID NO: 3), or the polynucleotide sequence may be codon optimized for expression in Lactococcus lactis (eg, sequence Number 5).

  The term “host cell” in the context of the present invention includes any cell that contains either a polynucleotide sequence encoding HP as described herein or an expression vector comprising said polynucleotide sequence described herein. Host cells can be used for the recombinant production of proteins having specific properties as defined herein. The host cell may contain a heterologous polynucleotide sequence encoding HP, or may be a cell that expresses the native HP polynucleotide. For example, the host cell may be from a Bacteroides species, such as Bacteroides tetaiotaomicron.

  The term “host cell” as used herein may be used synonymously with “host organism” and “host microorganism”.

  Accordingly, a host cell transformed with or transfected with a polynucleotide sequence encoding HP as described herein or transformed with an expression vector comprising the polynucleotide sequence described herein. Host cells transformed or transfected with the expression vectors are provided.

  The term “transfected cell” or “transfected host cell” as used herein includes, or is used herein, to include a polynucleotide sequence encoding the HP described herein. A host cell that has been transfected to contain an expression vector comprising the polynucleotide sequence described. In addition, or alternatively, the host cell has been transformed with a nucleotide sequence that causes the polynucleotide sequence encoding HP to be overexpressed in the host cell. For example, a promoter is inserted into the genome of the host cell, thereby allowing the host cell to overexpress the endogenous polynucleotide sequence encoding HP.

  The term “transformed cell” or “transformed host cell” means a host cell having an altered genetic structure.

  The term “host cell” includes any cell that can be transfected or transduced with a vector.

  The host cell will be selected to be compatible with the vector and can be, for example, a prokaryotic (eg, bacterium), fungus, yeast or plant cell.

  A host cell comprising a polynucleotide sequence encoding the polypeptide HP or a host cell comprising an expression vector comprising the polynucleotide sequence can be used to express the polypeptide HP under in vitro, in vivo and ex vivo conditions. is expected.

  In one embodiment, the host cell is a microorganism such as a bacterium. Typically, a microorganism that inhabits the digestive tract or part of the digestive tract. Examples of suitable bacterial host cells are gram positive or gram negative bacterial species. For example, the host cell may be Bacteroides spp. (Eg Bacteroides tetaiotaomicron), E. coli, Lactococcus spp (eg L. lactis), Lactobacillus spp, Bifidobacterium It is expected that it can be selected from the group consisting of genus species (Bifidobacterium spp), and Streptococcus spp (eg, Streptococcus thermophilus).

  In one embodiment, the host cell comprises an exogenous polynucleotide sequence encoding HP.

  In another embodiment, the host cell comprises an endogenous polynucleotide sequence encoding HP. For example, an endogenous polynucleotide sequence that is under the control of a non-native promoter (eg, a constitutive promoter). In a further example, the host cell contains multiple copies of the endogenous polynucleotide sequence.

  The term “host cell” also does not include native nucleotide coding sequences in their native environment when under the control of the native promoter of the coding sequence in its native environment. An example of a host cell having a native nucleotide sequence that is not in its own natural environment is Bacteroides tetaiotaomicron VPI-5482, which comprises SEQ ID NO: 1 under the control of a non-native promoter (eg, a constitutive promoter). In another example, the Bacteroides tetaiotaomicron VPI-5482 comprising SEQ ID NO: 1 has multiple copies of SEQ ID NO: 1.

  Depending on the nature of the nucleotide sequence and / or desirability of further processing of the expressed protein, eukaryotic hosts such as yeast or other fungi or insect cells (eg, insect Sf9 cells) may be used. In general, yeast cells are preferred over fungal cells because they are easier to manipulate. However, some proteins are not fully secreted from yeast cells or in some cases are not properly processed (eg, excess glycosylation in yeast). In these cases, another fungal host organism should be selected.

  The use of suitable host cells, such as yeast, fungal and plant host cells, may optionally require post-translational modifications (e.g., to impart optimal biological activity to the polypeptides described herein (e.g., It is expected that it can be prepared for myristic acid addition, glycosylation, shortening, lipidation and tyrosine, serine or threonine phosphorylation).

  It is expected that the host cells can be cultured under suitable conditions that allow expression of the polypeptide.

  In some embodiments, the polypeptide can be extracted from the host cell by various techniques known in the art, including enzymatic, chemical and / or osmotic lysis and physical degradation. It is expected that the polypeptide can be purified and isolated in a manner known per se.

Transformation of host cells Techniques for transformation of prokaryotic hosts are well described in the art, for example Sambrook et al (Molecular Cloning: A Laboratory Manual, 2nd edition, 1989, Cold Spring Harbor Laboratory Press). Please refer to. If a prokaryotic host is used, it may be necessary to appropriately modify the nucleotide sequence prior to transformation, such as by removing introns.

  Filamentous fungal cells can be transformed using various methods known in the art, for example, methods involving protoplast formation and protoplast transformation in a known manner, followed by cell wall regeneration. is expected. The use of Aspergillus as a host microorganism is described in EP0238023.

  Another host organism may be a plant. An overview of the general techniques used to transform plants is given by Potrykus (Annu Rev Plant Physiol Plant Mol Biol [1991] 42: 205-225) and Christou (Agro-Food-Industry Hi-Tech). March / April 1994 17-27). Further teachings regarding plant transformation are expected to be found in EP 0449375.

  General teachings on fungal, yeast and plant transformation are presented in the sections below.

Transformed fungus The host cell may be a fungus, such as a mold. Examples of suitable such hosts include any member belonging to the genus Thermomyces, Acremonium, Aspergillus, Aochobi, Astragalus, Acapanga, Trichoderma and the like.

  In one embodiment, the host cell may be a filamentous fungus.

  Filamentous transformation is discussed in US Pat. No. 5,741,665, and standard techniques for filamentous fungal transformation and its fungal culture are stated to be well known in the art. A broad review of techniques such as applied to N. crassa can be found, for example, in Davis and de Serres, Methods Enzymol (1971) 17A: 79-143.

  Further teachings that can also be utilized for transformation of filamentous fungi are outlined in US Pat. No. 5,674,707.

  In addition, gene expression in filamentous fungi is taught in Punt et al. (2002) Trends Biotechnol 2002 May; 20 (5): 200-6, Archer & Peberdy Crit Rev Biotechnol (1997) 17 (4): 273-306 Has been.

  The present specification encompasses the production of transgenic filamentous fungi according to the present specification prepared by use of these standard techniques.

  In one embodiment, the host organism can be of the genus Aspergillus, such as Aspergillus niger.

  For example, Turner G. 1994 (Vectors for genetic manipulation. In: Martinelli SD, Kinghorn JR (Editors) Aspergillus: 50 years on. Progress in industrial microbiology vol 29. Elsevier Amsterdam 1994. pp. 641-666) can also be prepared.

Transformed Yeast In another embodiment, the transgenic organism can be a yeast.

  A review of the principles of heterologous gene expression in yeast is provided, for example, in Methods Mol Biol (1995), 49: 341-54, and Curr Opin Biotechnol (1997) Oct; 8 (5): 554-60.

  In this context, yeasts, such as Saccharomyces cerevisi or Pichia pastoris species (see FEMS Microbiol Rev (see 2000 24 (1): 45-66), are vehicles for heterologous gene expression. May be used as

  An overview of the principles of heterologous gene expression and gene product secretion in budding yeast can be found in E Hinchcliffe E Kenny (1993, "Yeast as a vehicle for the expression of heterologous genes", Yeasts, Vol 5, Anthony H Rose and J Stuart Harrison eds, 2nd edition, Academic Press).

  Several transformation procedures have been developed for yeast transformation. For example, transgenic Saccharomyces according to the present invention is described by Hinnnen et al. (1978, Proceedings of the National Academy of Sciences of the USA 75, 1929), Beggs, JD (1978, Nature, London, 275, 104) and It can be prepared according to the teachings of Ito, H et al (1983, J Bacteriology 153, 163-168).

  It is expected that transformed yeast cells can be selected using a variety of selectable markers, such as auxotrophic markers, dominant antibiotic resistance markers.

Transformed Plant / Plant Cell A suitable host cell for the present invention may be a plant cell. In this regard, the basic principle of construction of genetically modified plants is to insert genetic information into the plant genome to achieve stable maintenance of the inserted genetic material. General techniques are reviewed in Potrykus (Annu Rev Plant Physiol Plant Mol Biol [1991] 42: 205-225) and Christou (Agro-Food-Industry Hi-Tech March / April 1994 17-27). Expected to be found.

  Direct infection of plant tissue by Agrobacterium is a simple technique that has been widely used, butcher DNet al., (1980), Tissue Culture Methods for Plant Pathologists, eds .: DS Ingrams and JP Helgeson , 203-208.

  Other techniques for transforming plants include ballistic transformation, silicon carbide whisker technology (Frame BR, Drayton PR, Bagnaall SV, Lewnau CJ, Bullock WP, Wilson HM, Dunwell JM, Thompson JA & Wang K ( 1994) Production of fertile transgenic maize plants by silicon carbide whisker-mediated transformation, See The Plant Journal 6: 941-948) and virus transformation techniques (eg Meyer P, Heidmann I & Niedenhof I (1992) The use of cassava mosaic virus as a vector system for plants, Gene 110: 213-217).

  Further teachings regarding plant transformation are expected to be found in EP 0449375.

  It is expected that plant cells can be grown and maintained according to well-known tissue culture methods, for example, by culturing the cells in a suitable culture medium supplemented with necessary growth factors such as amino acids, plant hormones, vitamins, etc. The

  In a further aspect, the present description relates to a vector system that carries a nucleotide sequence or construct according to the present description and that can introduce the nucleotide sequence or construct into the genome of an organism such as a plant. A vector system may contain one vector but may contain two vectors. In the case of two vectors, the vector system is usually called a binary vector system. The binary vector system is described in further detail in Gynheung An et al., (1980), Binary Vectors, Plant Molecular Biology Manual A3, 1-19.

  One widely used system for plant cell transformation uses Ti plasmids from Agrobacterium tumefaciens or Ri plasmids from Agrobacterium rhizogenes. An et al., (1986), Plant Physiol. 81, 301-305 and Butcher D. N. et al., (1980), Tissue Culture Methods for Plant Pathologists, eds .: D.S. Ingrams and J.P. Helgeson, 203-208. Following the introduction of the desired promoter in the plant or each of the constructs or nucleotide sequences according to the invention, the presence and / or insertion of additional DNA sequences may be necessary. For example, when using plant cell Ti or Ri plasmids for transformation, bind at least the right border of Ti and Ri plasmid T-DNA, but often the left and right borders, as adjacent regions of the introduced gene. be able to. The use of T-DNA for transformation of plant cells has been thoroughly studied and is described in EP 0120516; Hoekema, in: The Binary Plant Vector System Offset-drukkerij Kanters BB, Alblasserdam, 1985, Chapter V; Fraley, et al., Crit. Rev. Plant Sci., 4: 1-46; and An et al., EMBO J. (1985) 4: 277-284.

Culture and Production Host cells transformed with the nucleotide sequences described herein are cultured under conditions that facilitate production of the encoded polypeptide and facilitate recovery of the polypeptide from the cell and / or culture medium. Can be done.

  The medium used for culturing the cells may be any conventional medium suitable for the growth of the host cells and the expression of the polypeptide.

  Proteins produced by recombinant cells may be displayed on the cell surface.

  Using well-known procedures, the protein may be secreted from the host cell and may be conveniently recovered from the culture medium.

Secretion In many cases, it is desirable for the protein to be secreted from the expression host cell into a culture medium where the protein can be more easily recovered. According to the present specification, it is expected that the secretory leader sequence can be selected based on the desired expression host. Hybrid signal sequences may be used in connection with the present invention.

  Typical examples of heterologous secretory leader sequences are those derived from the fungal amyloglucosidase (AG) gene (glaA, eg, both 18 and 24 amino acid versions from Aspergillus), a factor gene (eg, yeast, eg , Derived from Saccharomyces, Kluyveromyces and Hansenula, or derived from the α-amylase gene (gonococci).

  As an example, secretion of heterologous proteins in E. coli is reviewed in Methods Enzymol (1990) 182: 132-43.

Expression Vector The term “expression vector” means a construct capable of expression in vivo, ex vivo or in vitro.

  The term “construct” is synonymous with terms such as “conjugate”, “cassette” and “hybrid”, and may be directly or indirectly linked to a promoter. The nucleotide sequence described in An example of indirect binding is the provision of a spacer group intervening between the promoter and the nucleotide sequence of the invention, eg, an intron sequence such as a Sh1-intron or an ADH intron. The same is true for the term “fused” with respect to the present invention, including direct or indirect binding. In some cases, these terms include natural combinations of nucleotide sequences encoding proteins normally associated with wild-type gene promoters, both of which are in their natural environment. do not do.

  The construct may even contain or express a marker that allows selection of the gene construct.

  For some applications, the construct includes at least a nucleotide sequence described herein that is operably linked to a promoter.

  The nucleotide sequence herein may be present in a vector, wherein the nucleotide sequence is operably linked to a regulatory sequence capable of effecting expression of the nucleotide sequence by a suitable host cell. Yes.

  In some embodiments, the polynucleotide sequence encoding the polypeptide HP of the expression vector will or will be transformed with the polynucleotide sequence or the polynucleotide sequence will be transfected. It may be codon optimized for the host cell.

  The term “operably linked” refers to a juxtaposition that exists in a relationship that allows the described components to function in their intended manner. A regulatory sequence “operably linked” to a coding sequence is ligated in such a way that expression of the coding sequence is achieved under conditions compatible with the control sequences. For example, a promoter is operably linked to a coding sequence when it controls transcription of the coding sequence.

  The term “regulatory sequence” includes promoters and enhancers as well as other expression control signals.

  The term “promoter” is used in the normal sense of the art, eg an RNA polymerase binding site. The promoter may be heterologous to the nucleotide sequence or may be homologous.

  Enhanced expression of the nucleotide sequences described herein may be achieved by heterologous regulatory regions such as promoters, secretory leaders and termination regions.

  In one embodiment, the nucleotide sequence described herein is operably linked to at least one promoter.

  Other promoters may be used to direct the expression of the polypeptides described herein.

  Examples of suitable promoters for directing transcription of nucleotide sequences in bacterial, fungal or yeast hosts are well known in the art.

  A promoter can further include a mechanism for ensuring expression in a suitable host or for increasing expression. For example, the mechanism can be a storage area, such as a pre-Beno box or a TATA box.

  Once transformed into a suitable host, the vector can replicate and function independently of the host genome, or in some cases, can be integrated into the genome of a suitable host cell. In some cases, the term “integrated” encompasses stable integration into the genome.

  Vectors for use in the present invention may be transformed into a suitable host cell as described herein to effect expression of the polypeptides herein.

  The vector may be a plasmid, a phage particle, or simply a potential genomic insert.

  The selection of a vector, such as a plasmid, cosmid or phage vector, will often depend on the host cell into which it will be introduced.

  Vectors for use in the present invention may also contain one or more selectable marker genes, for example genes that confer antibiotic resistance, such as ampicillin, kanamycin, chloramphenicol or tetracycline resistance. is there. Alternatively, selection may be accomplished by cotransfection (as described in WO 91/17243).

  Vectors may be used in vitro, for example, for the production of RNA, or to transfect host cells, to transfect host cells, to transfect host cells, or to transfect host cells. Sometimes used to infect cells.

  The vector may further comprise a nucleotide sequence that allows the vector to replicate in the host cell. Examples of such sequences are the origins of replication of plasmids pUC19, pACYC177, pUB110, pE194, pAMB1 and pIJ702.

  In one embodiment, the expression vector comprises one or more polynucleotide sequences according to the present invention. The polynucleotide sequence may be heterologous to or homologous to the host cell transformed with or transfected with the expression vector.

Disorder Polypeptide HP, polynucleotide sequence encoding polypeptide HP, host cell comprising said polynucleotide sequence, or host cell comprising an expression vector comprising said polynucleotide sequence are used for the treatment and / or prevention of a disorder of interest. The disorder can be an inflammatory disorder and / or an autoimmune disorder. The disorder may be a disorder of the CNS, including autism.

  In one embodiment, the disorder is a gastrointestinal tract, a portion of the gastrointestinal tract, liver, hepatocytes, epithelial cells, epidermal cells, nerve cells, pancreas and / or pancreatic cells (eg, islets of Langerhans), kidney, spleen, lung and heart. And / or affect those cells.

  Examples of portions (ie, portions) of the digestive tract include the mouth, esophagus, stomach and intestine (eg, small intestine (eg, duodenum, jejunum and ileum) and / or large intestine (eg, cecum, ascending colon, transverse colon, descending colon and Sigmoid colon)).

  Examples of epithelial cells include intestinal, oral, lung, nasal and vaginal epithelial cells.

  In one embodiment, the disorder is inflammatory bowel disorder (IBD), colitis, rheumatoid arthritis, psoriasis, multiple sclerosis, type I diabetes, celiac disease, atopic dermatitis, rhinitis, irritable bowel Syndrome (IBS, irritable bowel syndrome), ulcerative colitis, ileal cystitis, Crohn's disease, functional dyspepsia, atopic disease, necrotizing enterocolitis, nonalcoholic fatty liver disease, gastrointestinal infection, lupus, nephritis / glomerulonephritis , Selected from the group consisting of asthma, COPD, myocarditis and combinations thereof.

  In one aspect, the disorder affects the intestine.

  In one aspect, the disorder is an inflammatory disorder. For example, the disorder is an inflammatory bowel disorder (IBD) such as Crohn's disease.

  In one aspect, the disorder is an autoimmune disorder. For example, autoimmune disorders include ulcerative colitis, ileal cystitis, rheumatoid arthritis, psoriasis, multiple sclerosis, type I diabetes, allergy (including celiac disease), atopic dermatitis, rhinitis, lupus, nephritis / glomera Selected from the group consisting of nephritis, asthma, COPD and myocarditis.

Subjects In one embodiment, the subject is a monogastric animal.

  Examples of monogastric animals include poultry, humans, rats, pigs, dogs, cats, horses and rabbits.

  In another embodiment, the subject is a mammal, such as a monogastric mammal.

  Examples of monogastric mammals include omnivores (eg, humans, rats and pigs), carnivores (eg, dogs and cats), and herbivores (eg, horses and rabbits).

  In one embodiment, the subject is a human.

  In one aspect, the subject has inflammatory bowel disorder (IBD), colitis, rheumatoid arthritis, psoriasis, multiple sclerosis, type I diabetes, celiac disease, atopic dermatitis, rhinitis, irritable bowel syndrome (IBS), Ulcerative colitis, ileal cystitis, Crohn's disease, functional dyspepsia, atopic disease, necrotizing enterocolitis, nonalcoholic fatty liver disease, gastrointestinal infection, lupus, nephritis / glomerulonephritis, asthma, COPD, myocarditis and these Having a disorder selected from the group consisting of: For example, the subject has IBD.

Modulation / Adjustment The terms “modulation” and “modulation” are sometimes used synonymously herein.

  In one embodiment, the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence is a cell, tissue or organ of interest. Used to modulate inflammation.

  In one embodiment, the term “modulation” refers to increase and / or induction and / or promotion and / or activation. In an alternative embodiment, the term “modulation” refers to reduction and / or reduction and / or inhibition.

  In one embodiment, the term “modulation” refers to upregulation. In an alternative embodiment, the term “adjustment” refers to downward adjustment.

  In one embodiment, a polypeptide HP, a polynucleotide sequence encoding a polypeptide HP, or a host cell described herein reduces inflammation of a cell, tissue or organ. For example, the digestive tract, a portion (ie, part) of the digestive tract (ie, the intestine), liver, hepatocyte, epithelial cell, epidermal cell, nerve cell, pancreas and / or pancreatic cell (eg, islets of Langerhans), kidney, Reduce inflammation of the spleen, lungs and heart and / or their cells.

  In one example, inflammation of the gastrointestinal tract or part thereof (eg, intestine) is reduced.

  In another example, inflammation due to epithelial cells of a tissue or organ is reduced.

  The term “inflammation” as used herein refers to one or more of the following: redness, swelling, pain, tenderness, fever resulting from an inflammatory process induced by an overreaction of the immune system. And cell, tissue or organ dysfunction.

  In one embodiment, the number of inflamed cells of the subject encodes a polypeptide HP, or HP, as described herein after administration of a polypeptide, polynucleotide, or host cell described herein. Or at least 10%, 20%, 30%, 40% or 50% less than the number of inflamed cells in the subject prior to administering the host cell to the subject.

  In one embodiment, the amount of the inflamed tissue or organ of the subject is the amount of polypeptide HP, HP, as described herein after administration of the polypeptide, polynucleotide or host cell described herein. At least 10%, 20%, 30%, 40% or 50% less compared to the amount of inflamed tissue or organ of the subject prior to administration of the encoding polynucleotide sequence or host cell to the subject.

  In one embodiment, a polypeptide HP, a polynucleotide sequence encoding a polypeptide HP, or a host cell as described herein reduces inflammation due to epithelial cells of a tissue or organ.

  For example, an epithelial cell is an epithelial cell of the digestive tract or a part thereof (eg, intestine).

  Without wishing to be bound by theory, the polypeptide HP, polynucleotide sequence encoding the polypeptide HP, or host cell described herein may be a T cell in a subject (eg, referred to as a Treg). Increase the production of certain regulatory T cells). This increase in Treg number can counteract the effects of other effector T cells (also called Teff), such as Th1, Th17 and Th2, leading to inflammation, autoimmunity and allergic / atopic conditions. In Crohn's disease and ulcerative colitis, the Teff / Treg cell balance is lost.

  In one embodiment, the production of T cells in the subject comprises polypeptide HP, or a polynucleotide sequence encoding HP, as described herein after administration of a polypeptide, polynucleotide or host cell described herein. Or at least 10%, 20%, 30%, 40% or 50% more T cells, or more than 100% more T cells compared to the number of T cells in the subject prior to administering the host cell to the subject Is increased to exist.

Intestinal Barrier Integrity In one embodiment, a polypeptide HP, a polynucleotide sequence encoding a polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence is a subject intestine. Used to improve barrier integrity.

  The term “improving intestinal barrier integrity”, as used herein, means that the number and / or type of microorganisms that are transmitted from the subject's intestine to other cells is the polypeptide, polypeptide, or polypeptide described herein. After administration of a nucleotide or host cell, reduced compared to the number and / or type of microorganisms transmitted from the subject's intestine to other cells prior to administration of the polypeptide, polynucleotide or host cell described herein. Refers to that.

  In one embodiment, the number of microorganisms that propagate from the subject's intestine to other cells is determined by administration of a polypeptide, polynucleotide, or host cell described herein, polypeptide HP, or A polynucleotide sequence encoding HP, or at least 10%, 20%, 30%, 40% or 50 compared to the number of microorganisms transmitted from the subject's intestine to other cells prior to administration of the host cell to the subject. %Few.

  In one embodiment, the polypeptide HP, HP described herein is encoded for a microorganism that is transmitted from the intestine of the subject to other cells after administration of the polypeptide, polynucleotide or host cell described herein. There are at least 5%, 10%, 15%, or 20% less species compared to the polynucleotide sequence to be transmitted, or the type of microorganism that propagates from the subject's intestine to other cells prior to administering the host cell to the subject. .

Bacterial Level In one embodiment, a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence is a more beneficial microorganism. Used to modify the bacterial composition within a tissue or organ to provide a flora. For example, the present invention may be used to reduce the level of one or more types of lactose fermenting bacteria (eg, E. coli) in a subject tissue or organ and / or one or more in a subject tissue or organ. It can be used to reduce the level of two or more types of non-lactose fermenting bacteria.

  The term “reducing the level of one or more types of lactose-fermenting bacteria” as used herein describes the number of lactose-fermenting bacteria in a tissue or organ of interest as described herein. After administration of the polypeptide, polynucleotide or host cell, as compared to the number of lactose-fermenting bacteria in the tissue or organ of interest prior to administration of the polypeptide, polynucleotide or host cell described herein Refers to that.

  In one embodiment, the number of lactose-fermenting bacteria in a subject tissue or organ is determined by administering a polypeptide HP, HP, as described herein after administration of a polypeptide, polynucleotide or host cell as described herein. At least 10%, 20%, 30%, 40% or 50% less than the number of lactose-fermenting bacteria in the tissue or organ of the subject prior to administering the encoding polynucleotide sequence or host cell to the subject.

  Examples of lactose-fermenting bacteria include E. coli, Enterobacter and Klebsiella.

  The term “reducing the level of one or more types of non-lactose-fermenting bacteria” as used herein indicates the number of non-lactose-fermenting bacteria in a tissue or organ of interest as defined herein. Refers to being reduced after administration of the polypeptide, polynucleotide or host cell, as compared to the number of non-lactose fermenting bacteria in the tissue or organ of interest prior to administration of the polypeptide, polynucleotide or host cell. .

  In one embodiment, the number of non-lactose-fermenting bacteria in the tissue or organ of interest is the polypeptide HP, HP, as described herein after administration of the polypeptide, polynucleotide or host cell described herein. Or at least 10%, 20%, 30%, 40% or 50% compared to the number of non-lactose fermenting bacteria in the tissue or organ of the subject prior to administering the host cell to the subject Few.

  Examples of non-lactose fermenting bacteria include bacteria such as Salmonella, Proteus species, Pseudomonas aeruginosa and Shigella.

  In one embodiment, the tissue or organ is selected from the group consisting of mesenteric lymph nodes, liver, pancreas, spleen and combinations thereof.

Appetite and / or body weight regulation In one embodiment, a polypeptide HP, a polynucleotide sequence encoding the polypeptide, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence comprises: Used to regulate the appetite (eg, food intake) of a subject (eg, a subject with IBD).

  As used herein, the term “modulate appetite” or “modulate appetite” can modulate (eg, increase or decrease) a subject's desire to eat food. Point to.

  In one embodiment, the term “modulate” or “modulation” refers to an increase in appetite (eg, food intake). In an alternative embodiment, the term “modulate” or “modulation” refers to a decrease in appetite (eg, food intake).

  For example, a polypeptide, polynucleotide or host cell described herein maintains or stimulates a subject's appetite.

  In one embodiment, a polypeptide HP, a polynucleotide sequence encoding said polypeptide, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence has a subject (eg, IBD) Used to adjust the subject's weight.

  In one embodiment, the term “modulate” or “modulation” refers to weight gain. In an alternative embodiment, the term “modulate” or “modulation” refers to weight loss.

  For example, a polypeptide, polynucleotide or host cell described herein maintains or increases a subject's weight.

  Without wishing to be bound by theory, the polypeptide HP described herein, a polynucleotide sequence encoding HP, or a host cell may contain a gene (eg, a satiety hormone) associated with appetite suppression. It exerts stimulating effects on the appetite of the subject by down-regulating the expression of the encoding gene. Agt, Cartpt, Cck, Cxcl12 and Gcg are examples of genes associated with appetite regulation, and one or more down regulation of these genes is associated with appetite suppression.

  Cholecystokinin (Cck) and glucagon (Gcg, glucagon) are examples of satiety hormones.

  In one aspect, a polypeptide HP, a polynucleotide sequence encoding HP, or a host cell, as described herein, is administered after the subject is administered a polypeptide, polynucleotide or host cell as described herein. As described herein, the polypeptide HP, the polynucleotide sequence encoding HP, or the host cell is consumed at least 5%, 10% or 15% more food compared to the subject prior to administration to the subject, Stimulates the subject's appetite. Additionally or alternatively, a polypeptide HP, a polynucleotide sequence encoding HP, or a host cell, as described herein, can be obtained from the first administration of a polypeptide, polynucleotide or host cell as described herein. After months, the subject's body weight is at least 2%, 5%, or 10 as compared to the subject prior to administering the polypeptide HP, a polynucleotide sequence encoding HP, or a host cell as described herein. Stimulates the subject's appetite to increase by%.

  In one embodiment, a polypeptide HP, a polynucleotide sequence encoding HP, or a host cell as described herein reduces cholecystokinin (Cck) and / or glucagon (Gcg) levels in a subject's blood. Let

  In one aspect, the polypeptide HP described herein is a polynucleotide sequence that encodes HP, or a host cell is capable of measuring cholecystokinin (Cck) and / or glucagon (Gcg) levels in the blood of a subject. After administration of a polypeptide, polynucleotide or host cell as described herein, compared to a subject before administration of the polypeptide HP, or polynucleotide sequence encoding HP, or host cell as described herein to a subject By at least 5%, 10%, 15% or 20%.

  In one embodiment, the polypeptide HP, a polynucleotide sequence encoding HP, or a host cell as described herein is an expression of a gene encoding cholecystokinin (Cck) in one or more cells of interest and / Or down-regulate the expression of the gene encoding glucagon (Gcg).

  In one aspect, a polypeptide HP, a polynucleotide sequence encoding HP, or a host cell, as described herein, expresses expression of a gene encoding cholecystokinin (Cck) at its expression level (eg, mRNA level). ) After administration of the polypeptide, polynucleotide or host cell described herein, before administration of the polypeptide HP or polynucleotide sequence encoding HP or host cell described herein to the subject. Decreased to at least 5%, 10%, 15% or 20% lower than the expression level in the subject.

  In one aspect, a polypeptide HP, a polynucleotide sequence encoding HP, or a host cell as described herein has an expression level of a gene encoding glucagon (Gcg) whose expression level (eg, mRNA level) is In the subject after administration of the polypeptide, polynucleotide or host cell described herein, prior to administration of the polypeptide HP, polynucleotide sequence encoding HP, or host cell described herein, to the subject. Decrease by at least 5%, 10%, 15% or 20% lower than the expression level.

Maintaining the length of a portion of the intestine In one embodiment, a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence Is used to maintain the length of a portion of the subject's intestine (eg, the large and / or small intestine).

  Examples of intestinal parts (ie parts) include the small intestine (eg duodenum, jejunum and ileum) and / or the large intestine (eg cecum, ascending colon, transverse colon, descending colon and sigmoid colon).

  The term “maintain length” as used herein is described herein for the length of a portion of the intestine after administration of a polypeptide, polynucleotide or host cell described herein. Or no change compared to the length of that portion of the intestine prior to administration of the polypeptide, polynucleotide or host cell.

  In one embodiment, a polypeptide, polynucleotide sequence or host cell described herein prevents a reduction in colon length. In addition or alternatively, the polypeptides, polynucleotide sequences or host cells described herein prevent an increase in the length of the small intestine.

  In one embodiment, the small change in the length of the subject's large intestine is the length of the subject's large intestine prior to administration of the polypeptide HP, a polynucleotide sequence encoding HP, or a host cell as described herein. As compared to less than 5%, 2% or 1% of the length after administration of a polypeptide, polynucleotide or host cell described herein.

  In one embodiment, the small change in the length of the subject's small intestine is a polypeptide HP, a polynucleotide sequence encoding HP, or a length of the subject's small intestine prior to administration of the host cell to the subject. And an increase in length of less than 1%, 2%, 5%, 7% or 10% after administration of a polypeptide, polynucleotide or host cell described herein.

Intestinal disturbances In one embodiment, a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence is a subject (e.g., Used to reduce disturbances in the intestine (eg, large intestine) of a subject with IBD.

  The term “intestinal disturbance of the subject” as used herein refers to the effect on the integrity of the mucosal epithelium and / or the number of epithelial goblet cells and / or the number of immune cells infiltrating the lamina propria. Refers to impact.

  In one embodiment, the polypeptides, polynucleotide sequences or host cells herein reduce or prevent mucosal epithelial integrity disruption and / or reduce or prevent reduction in the number of epithelial goblet cells. And / or reduce or prevent immune cell infiltration into the lamina propria.

  In one embodiment, the reduction in disruption of mucosal epithelial integrity is from the intestinal lumen of a subject prior to administration of a polypeptide HP, a polynucleotide sequence encoding HP, or a host cell as described herein to the subject. At least 5%, 10%, 15 of the number of bacteria that migrate from the intestinal lumen to the intestinal cells after administration of the polypeptide, polynucleotide or host cell described herein, compared to the number of bacteria that migrate to the enterocytes % Or 20% reduction.

  In one embodiment, the increase in the number of epithelial goblet cells is a polypeptide HP, a polynucleotide sequence encoding HP, or a goblet cell of the subject's epithelium prior to administration to the subject, as described herein. An increase of at least 2%, 5%, 10%, 15% or 20% of the number of goblet cells of the subject's epithelium after administration of a polypeptide, polynucleotide or host cell described herein compared to the number It is.

  In one embodiment, the reduction of infiltration of immune cells into the lamina propria is over a period of time (eg, 24 hours), as described herein, polypeptide HP, a polynucleotide sequence encoding HP, or a host cell. The mucosa after administration of a polypeptide, polynucleotide or host cell described herein as compared to the number of immune cells (eg, T cells) that migrate into the lamina propria cells of the subject prior to administration to the subject A reduction, such as a reduction of at least 5%, 10%, 15%, 20% or 30% of the number of immune cells (eg, T cells) that migrate into the lamina propria.

Inflammatory and barrier integrity genes In one embodiment, a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence Is used to modulate the expression of one or more pro-inflammatory and / or anti-inflammatory genes and / or one or more barrier integrity genes in one or more cells of a subject Is done.

  In one embodiment, the term “modulate” refers to upregulation of the expression of one or more pro-inflammatory or anti-inflammatory genes. In an alternative embodiment, the term “modulate” refers to downregulation of the expression of one or more pro-inflammatory or anti-inflammatory genes.

  In one embodiment, a polypeptide HP, a polynucleotide sequence encoding a polypeptide HP, or a host cell as described herein is one or more pro-inflammatory genes in one or more cells of interest. Down-regulate the expression of.

  The term “pro-inflammatory gene” as used herein refers to a gene that, when expressed, promotes inflammation. Examples of pro-inflammatory genes include, but are not limited to IL1-β, IL4, IL5, IL6, IL8, IL12, IL13, IL17, IL21, IL22, IL23, IL27, IFNγ, CCL2, CCL3, CCL5, CCL20, CXCL5 , Genes encoding CXCL10, CXCL12, CXCL13 and TNF-α.

  In one embodiment, the pro-inflammatory gene is selected from the group consisting of IL6, CXCL10 and TNF-α.

  In one embodiment, the level of expression (eg, mRNA level) of one or more pro-inflammatory genes is such that the level is determined after administration of a polypeptide, polynucleotide or host cell described herein. At least 10%, 20%, 30%, 40% of the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, or the level of the subject prior to administering the host cell to the subject, as described herein Or reduced to 50% lower (ie down-regulated).

  The term “barrier integrity gene” as used herein refers to a gene that, when expressed, is involved in intestinal barrier functions such as barrier repair and prevention of microbial barrier crossing. Examples of barrier integrity genes include genes encoding Retlg | Retnlb, Si, Defa24, Hsd11b2, Hsd17b2, and Nr1d1 | Thra.

  In one embodiment, the term “modulate” refers to upregulation of the expression of one or more barrier integrity genes. In an alternative embodiment, the term “modulate” refers to downregulation of the expression of one or more barrier integrity genes.

  In one embodiment, a polypeptide HP, a polynucleotide sequence encoding a polypeptide HP, or a host cell described herein upregulates the expression of a barrier integrity gene in one or more cells of interest.

  In one embodiment, the barrier integrity gene is selected from the group consisting of Retlg | Retnlb, Si, Defa24, Hsd11b2, Hsd17b2 and Nr1d1 | Thra.

  In one embodiment, the expression level (eg, mRNA level) of one or more barrier integrity genes is such that the level is greater than after administration of a polypeptide, polynucleotide or host cell described herein. At least 10%, 20%, 30%, 40, as compared to the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, or its level prior to administration of the host cell to the subject, as described in the specification. % Or 50% higher (ie up-regulated).

  In one embodiment, polypeptide HP, a polynucleotide sequence encoding polypeptide HP, or a host cell described herein upregulates expression of an anti-inflammatory gene.

In one embodiment, a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence is a subject (eg, , Subjects with IBD) regenerated islet-derived 3 beta gene (Reg3b), resistin-like gamma-resistin-like beta gene (Retnlg | Retnlb), sucrase-isomaltase (alpha-glucosidase) gene (Si), Defensin alpha 24 gene (Defa24), hydroxysteroid 11-beta dehydrogenase 2 gene (Hsd11b2), hydroxysteroid (17-beta) dehydrogenase 2 gene (Hsd17b2), resist Is used to regulate the expression of one or more genes selected from the group consisting of the gene-like molecule-beta (RELMb) and the nuclear receptor 1D1 thyroid hormone receptor alpha gene (Nr1d1 | Thra) .

  The terms “Reg”, “Reg3”, and “Reg3b” as used herein are interchangeable.

  The terms “Hsd”, “Hsd17b2” or “Hsd17b2” as used herein are interchangeable.

  In one embodiment, the term “modulate” refers to upregulation of gene expression. In an alternative embodiment, the term “modulate” refers to down-regulation of gene expression.

  The present invention is useful for modulating the expression of pro-inflammatory and / or barrier integrity genes.

  To avoid misunderstanding, pro-inflammatory genes are IL-β, IL4, IL5, IL6, IL8, IL12, IL13, IL17, IL21, IL22, IL23, IL27, IFNα, CCL2, CCL3, CCL5, CCL20, CXCL5, Includes CXCL10, CXCL12, CXCL13 and TNF-α.

  To avoid misunderstandings, barrier integrity genes include Retlnlg / Retnlb, Si, Defa24, Hsd11b2, Hsd17b2, and Nrd1 \ Thra.

  In one embodiment, the polypeptide, polynucleotide sequence or host cell described herein comprises a regenerated islet-derived 3 beta gene (Reg3b), a resistin-like gamma resistin-like beta gene (Retlnlg | Retnlb), resistin-like molecule-beta (RELMb), the expression of one or more genes selected from the group consisting of a sucrase-isomaltase (alpha-glucosidase) gene (Si), and a defensin alpha24 gene (Defa24). For example, the expression level (eg, mRNA level) of one or more genes selected from the group can be determined after the administration of a polypeptide, polynucleotide or host cell described herein. At least 10%, 20%, 30%, 40, as compared to the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, or its level prior to administration of the host cell to the subject, as described in the specification. % Or 50% lower (ie down adjusted).

  In one embodiment, the polypeptide, polynucleotide sequence or host cell described herein comprises a hydroxysteroid 11-beta dehydrogenase 2 gene (Hsd11b2), a hydroxysteroid (17-beta) dehydrogenase 2 gene (Hsd17b2), and a nucleus Increases the expression of one or more genes selected from the group consisting of the internal receptor 1D1 thyroid hormone receptor alpha gene (Nr1d1 | Thra). For example, the expression level (eg, mRNA level) of one or more genes selected from the group can be determined after the administration of a polypeptide, polynucleotide or host cell described herein. At least 10%, 20%, 30%, 40% of the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, or the level of the subject prior to administering the host cell to the subject, as described herein Or increased to 50% higher (ie up-regulated).

In one embodiment, the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence is a subject 1 Or used to reduce the activation of pro-inflammatory pathways in multiple cells.

  Reduction in activation of a pro-inflammatory pathway can be determined by determining the inflammation of the subject.

  Inflammation in a subject is determined by determining the levels of pro-inflammatory cytokines and chemokines in the subject's tissues, serum and / or stool samples before and after administration of the polypeptides, polynucleotides or host cells described herein to the subject. This can be determined. For example, the level of one or more of the following can be monitored: IL-1, IL-4, IL5, IL6, IL-8, IL-12, IL-13, IL-17, IL -21, IL-22, IL23, TNFα, IFNγ, CXCL1, CXCL10, CCL20 serum and fecal calprotectin, SA1009 / SA1008 calcium binding protein, and type 1 interferons, CD markers such as CD163, CD14, inflammatory transcription factors For example, NF-κβ, STAT and MAP kinases, c-reactive protein (CRP), erythrocyte sedimentation rate (ESR), complement protein, serum albumin, target tissue and organ histology Evaluation, disease activity index.

  In one embodiment, the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence is one or more cells of interest It is used to reduce the activity and / or expression of NF-κβ in (eg epithelial cells, epidermal cells, nerve cells, liver, spleen, kidney, lung, heart and / or pancreatic cells).

  For example, the activity of NF-κβ can be determined after administration of a polypeptide, polynucleotide or host cell described herein, wherein the polypeptide HP is a polynucleotide sequence or host cell encoding the polypeptide HP. Is reduced by at least 10%, 20%, 30%, 40% or 50% compared to its level in the subject prior to administration.

  For example, the expression level (eg, mRNA) of NF-κβ is such that, after administration of a polypeptide, polynucleotide or host cell as described herein, the level of polypeptide HP, polypeptide as described herein. A polynucleotide sequence encoding HP or reduced to at least 10%, 20%, 30%, 40% or 50% lower than that level of the subject prior to administration to the subject (ie, Adjusted downward).

Gastrointestinal tract The gastrointestinal tract includes the mouth, esophagus, stomach and intestines (eg, small intestine (eg, duodenum, jejunum and ileum) and / or large intestine (eg, cecum, ascending colon, transverse colon, descending colon and sigmoid colon).

  In this specification, the term “colon” may be used synonymously with the term “colon”.

  In one embodiment, the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence is responsible for the health of the subject's digestive tract. Used to improve.

  The term “improving gastrointestinal health” as used herein refers to reducing the level of inflammation in the gastrointestinal tract or part thereof and / or improving the gut microbiota.

  In one embodiment, the level of inflammation in the gastrointestinal tract is administered to a subject with a polypeptide, polynucleotide or host cell described herein after administration of the polypeptide, polynucleotide or host cell described herein. At least 10%, 20%, 30%, 40% or 50% lower than the level of inflammation in the digestive tract of the previous subject.

  In one embodiment, the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence improves the gut microbiota of the subject Used to do.

  The term “intestinal microflora” as used herein refers to microorganisms that inhabit the digestive tract of a host animal. These microorganisms perform a wide variety of metabolic functions, structural functions, protective functions, and other beneficial functions.

  As used herein, the term “improving gut microbiota” increases the number and / or type of desirable microorganisms present in the intestine of a subject (eg, a host) and / or said desirable It refers to increasing activity in terms of microbial metabolic function, structural function, protective function and other beneficial functions. The term “improving intestinal microbiota” refers to reducing the number and / or type of undesirable microorganisms present in the gut of a subject (eg, host) and / or metabolic function, structure of said undesirable microorganisms. May also refer to reducing activity in terms of function, protective function and other beneficial functions.

  Desirable microorganisms in the intestine of the host are microorganisms that have protective and beneficial functions. Firmicutes and / or bacteroidetes bacteria are examples of desirable microorganisms in the intestine of the host.

  Undesirable microorganisms in the intestine of the host are microorganisms that can interfere with the metabolic, structural, protective and other beneficial functions of desirable microorganisms in the intestine that have protective and beneficial functions. In addition or alternatively, undesirable microorganisms are those that cause, for example, inflammation and / or diarrhea. E. coli (ETEC, EPEC, EIEC, EHEC and / or EAEC) are examples of undesirable microorganisms in the host intestine.

  For example, the number (ie, level) of the bacteria of the Fermicutes and / or Bacteroides is increased and the number of E. coli is decreased. Such an improvement of the intestinal microflora is achieved in a subject having inflammatory bowel disease (IBD), a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or This can occur when a host cell containing an expression vector comprising the polynucleotide sequence is administered to a subject.

  In one embodiment, the number of desirable microorganisms (eg, Pharmicutes and Bacteroides bacteria) present in the intestine of a subject (eg, host) is the number of microorganisms described herein, After administration of the polynucleotide or host cell, at least 10%, 20%, 30%, 40% or compared to the level in the subject prior to administering the polypeptide, polynucleotide or host cell described herein to the subject Increased to be 50% higher or higher than 100%. In addition or alternatively, the type of desirable microorganism (eg, a bacterium of Clostridium cluster XIVa) present in the gut of a subject (eg, host) is a polypeptide, polynucleotide or After administration of the host cell, there are at least 2%, 5%, 10% or 15% more microbial species compared to the type in the subject prior to administering the polypeptide, polynucleotide or host cell described herein to the subject. Increased to exist.

  In one embodiment, the protein of the invention modifies the bacterial composition of the subject's intestines to provide a beneficial microflora. For example, the number of undesirable microorganisms (eg, E. coli (ETEC, EPEC, EIEC, EHEC, and / or EAEC)) present in the intestine of a subject (eg, host) is the number of microorganisms described herein. After administration of the polypeptide, polynucleotide or host cell, at least 10%, 20%, 30% compared to the level in the subject prior to administering the polypeptide, polynucleotide or host cell described herein to the subject, Decrease to be 40% or 50% lower. In addition, or alternatively, the type of undesirable microorganism (eg, E. coli) present in the gut of the subject (eg, host) may be determined after administration of the polypeptide, polynucleotide or host cell described herein. There is an undesirable microbial species present that is at least 1%, 2%, 5%, or 10% less compared to the type in the subject prior to administering the polypeptide, polynucleotide or host cell described herein to the subject Reduced to

Encapsulation In one embodiment, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence is encapsulated.

  In a further embodiment, a pharmaceutical composition comprising a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence is encapsulated. .

  In another embodiment, a nutritional supplement comprising a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence is encapsulated. Yes.

  In further embodiments, the livestock feed, food, dietary supplement or food additive described herein is encapsulated.

  The term “encapsulated”, as used herein, protects a polypeptide, polynucleotide or host cell described herein from incompatible environments, and thus a polypeptide, polynucleotide or host cell. Refers to a means by which a polypeptide, polynucleotide or host cell can be delivered to a target site (eg, intestine) without degradation or significant degradation such that can affect the target site. Examples are enteric capsules or enteric resistant capsules.

  Even when the purpose of encapsulation is the isolation of a polypeptide, polynucleotide or host cell from its environment, the protective coating or tablet must be broken during the desired action. Breaking of the protective coating or tablet skin typically occurs by the application of chemical and physical stimuli such as pressure, enzyme attack, chemical reaction and physical disruption.

  For example, encapsulation can provide a polypeptide, polynucleotide, or host cell such that an amount of the polypeptide, polynucleotide or host cell that is effective to exert an effect at the target site can be delivered to the target site (eg, intestine). Allow ingestion of nucleotides or host cells.

Pharmaceutical composition In one embodiment, the pharmaceutical composition comprises a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence. A pharmaceutically acceptable excipient, carrier or diluent.

  The pharmaceutical composition may be any pharmaceutical composition. In one embodiment, the pharmaceutical composition can be administered orally, enterally or rectally. For example, the composition may be an edible composition. “Edible” means a substance that is approved for human or animal consumption.

  The pharmaceutical composition may be for human or animal use in human and veterinary medicine.

  Examples of such suitable excipients for various different forms of the pharmaceutical compositions described herein can be found in “Handbook of Pharmaceutical Excipients, 2nd Edition, (1994), Edited by A Wade and PJ Weller. Expected to be found.

  Carriers or diluents that are acceptable for therapeutic use are well known in the pharmaceutical art and are described, for example, in Remington's Pharmaceutical Sciences, Mack Publishing Co. (A. R. Gennaro edit. 1985).

  Examples of suitable carriers include lactose, starch, glucose, methylcellulose, magnesium stearate, mannitol, sorbitol and the like.

  Examples of suitable diluents include one or more of water, ethanol, glycerol, propylene glycol and glycerin, and combinations thereof.

  The choice of pharmaceutical carrier, excipient or diluent can be selected with regard to the intended route of administration and standard pharmaceutical practice. The pharmaceutical composition comprises any suitable binder, lubricant, suspending agent, coating agent, solubilizer as a carrier, excipient or diluent or in addition to the carrier, excipient or diluent. May be included.

  Examples of suitable binders include starch, gelatin, natural sugars such as glucose, anhydrous lactose, free flowing lactose, beta-lactose, corn sweeteners, natural and synthetic gums such as gum arabic, tragacanth or sodium alginate, carboxy Examples include methyl cellulose and polyethylene glycol.

  Examples of suitable lubricants include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride and the like.

  Preservatives, stabilizers, dyes and even flavoring agents may be supplied to the pharmaceutical composition. Examples of preservatives include sodium benzoate, sorbic acid, and esters of p-hydroxybenzoic acid. Antioxidants and suspending agents may be used.

  In one embodiment, the polypeptide, polynucleotide sequence or host cell of the pharmaceutical composition is encapsulated.

  In another embodiment, the polypeptide of the pharmaceutical composition is a recombinant polypeptide.

  In further embodiments, the polynucleotide sequence of the pharmaceutical composition encodes a recombinant polypeptide.

  In another embodiment, the host cell of the pharmaceutical composition produces or is capable of producing a recombinant polypeptide.

  In a further embodiment, the expression vector comprises the polynucleotide sequence of the pharmaceutical composition.

  The medicament may be in the form of a solution or may be present as a solid, depending on the application and / or method of use and / or method of administration.

  As used herein, the term “drug” encompasses drugs for both human and veterinary use in human and veterinary medicine. In addition, the term “drug” as used herein means any substance that provides a therapeutic and / or beneficial effect. The term “drug”, as used herein, is not necessarily limited to substances that require marketing authorization, but is cosmetic, fortified food, food (eg, including feed and beverages), probiotic cultures, Substances that can be used in nutritional supplements and naturopathic drugs can be included. In addition, the term “drug” as used herein includes products designed for incorporation into animal feed, such as livestock feed and / or pet food.

Nutritional supplements Nutritionally acceptable carriers, diluents and excipients include those suitable for human or animal consumption and used as standard in the food industry. Typical nutritionally acceptable carriers, diluents and excipients will be familiar to those skilled in the art.

  In one embodiment, the nutritional supplement comprises a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence; And an acceptable excipient, carrier or diluent.

  In one example, the nutritional supplement polypeptide, polynucleotide sequence or host cell is encapsulated.

  In another example, the nutritional supplement polypeptide is a recombinant polypeptide.

  In a further embodiment, the polynucleotide sequence of the nutritional supplement encodes a recombinant polypeptide.

  In another embodiment, the nutritional supplement host cell produces or is capable of producing a recombinant polypeptide.

  In a further example, the nutritional supplement polynucleotide is included in an expression vector.

Livestock feed / product A further aspect of the invention includes a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, It relates to livestock feed, food, nutritional supplements and food additives.

  The terms “livestock feed”, “food”, “food additive” and “dietary supplement” as used herein are meant to encompass any ingestible product, which may be a solid, jelly or liquid. It is intended.

  The term “food” is used in a broad sense and encompasses animal food (ie, feed) as well as human food. In one aspect, the food product is for human consumption. Examples of food products include dairy products (eg, milk, cheese, beverages containing whey protein, milk drinks, lactic acid bacteria drinks, yogurt, drinking yogurt), bakery products, beverages, and powdered beverages.

  The “livestock feed”, “food”, “food additive” and “nutritional supplement” may be in the form of a solution or a solid depending on the use and / or method of use and / or method of administration It may exist as

  As used herein, the term “nutritional supplement” includes formulations that are or can be added as a nutritional supplement to food or livestock feed. The term “nutritional supplement” as used herein requires gelling, texture-imparting, stabilizing, suspending, film-forming and structuring, maintaining juiciness, and improving texture. Also refers to formulations that can be used at low levels in a wide variety of products without increasing viscosity.

  Examples of suitable foods include functional foods, food compositions, pet foods, livestock feeds, health foods, livestock feeds and the like. In one aspect, the food is a health food.

  As used herein, the term “functional food” means a food that not only can provide a nutritional effect, but can also provide additional beneficial effects to the consumer. Thus, a functional food is supplemented with ingredients or ingredients (e.g., those described herein) that impart a specific function, such as a medical or physiological benefit, to the food other than just a nutritional effect, It is an ordinary food.

  Examples of specific foods that can be used in the present invention include milk-based products for humans, ready-to-eat desserts, such as powders for reconstitution with milk or water, chocolate milk drinks, malt drinks, as is Or a ready-to-eat dish, an instant dish or drink, or a food composition corresponding to a full or partial meal for pets or livestock.

  In one aspect, the livestock feed, food, dietary supplement or food additive according to the invention is intended for humans, pets or livestock, eg monogastric animals. The livestock feed, food, dietary supplement or food additive may be for an animal selected from the group consisting of dogs, cats, pigs, horses or poultry. In a further embodiment, the food, dietary supplement or food additive is for adult species, particularly for adults.

  The term “milk-based product” as used herein means a liquid or semi-solid milk or whey-based product with varying fat content. Milk-based products include, for example, milk, goat milk, sheep milk, skim milk, whole milk, reduced milk from powdered milk, and whey that has not been processed at all, or processed products such as yogurt, curd, curd, acid milk Acidic whole milk, buttermilk and other acidic dairy products. Another important group includes milk drinks such as whey drinks, fermented milk, condensed milk, infant or infant milk, flavored milk, ice cream, milk-containing foods such as sweet confectionery.

  The livestock feed, food, dietary supplement or food additive of the present invention may be a food supplement or may be added to a food supplement, and the food supplement is herein referred to as a dietary supplement, nutritional supplement or Also called food additive.

  Livestock feeds, foodstuffs, dietary supplements or food additives according to the invention may be used for animal nutrition (eg for pig nutrition), especially in early weaning and growing fattening periods. Livestock feeds, foods, dietary supplements or food additives can enhance immune function, reduce and prevent infectious diseases, beneficially change microbiota composition, and increase animal growth and Expected to improve capacity.

  In one embodiment, the livestock feed, food, dietary supplement or food additive is encapsulated.

  In one embodiment, the livestock feed, food, dietary supplement or food additive polypeptide is a recombinant polypeptide.

  In one example, a livestock feed, food, dietary supplement or food additive polynucleotide is included in the expression vector.

Administration The pharmaceutical composition, nutritional supplement, livestock feed, food, dietary supplement or food additive of the present invention is oral, rectal, vaginal, parenteral, intramuscular, intraperitoneal, intraarterial, intrathecal, intrabronchial Applicable to subcutaneous, intradermal, intravenous, nasal, buccal or sublingual administration routes.

  In one aspect, the pharmaceutical composition, nutritional supplement, livestock feed, food, dietary supplement or food additive of the present invention is adapted for oral, rectal, vaginal, parenteral, nasal, buccal or sublingual administration routes. .

  In a further aspect, the pharmaceutical composition, nutritional supplement, livestock feed, food, dietary supplement or food additive of the present invention is adapted for oral administration.

  For oral administration, particular use is made of compressed tablets, pills, tablets, gellules, drops and capsules.

  Other dosage forms include solutions or emulsions that can be injected intravenously, intraarterially, intrathecally, subcutaneously, intradermally, intraperitoneally or intramuscularly and prepared from sterile or sterilizable solutions. The pharmaceutical compositions of the present invention may be in the form of suppositories, pessaries, suspensions, emulsions, lotions, ointments, creams, gels, sprays, solutions or powders.

  An alternative to transdermal administration is through the use of skin patches. For example, the active ingredient can be added to a cream made of an aqueous emulsion of polyethylene glycol or liquid paraffin. In another example, the active ingredients can be incorporated into ointments composed of white wax or white petrolatum base together with the stabilizers and preservatives described above.

  A pharmaceutical composition, nutritional supplement, livestock feed, food, dietary supplement or food additive in unit dosage form, i.e. in the form of a unit dose, or in individual parts containing multiple or small units of a unit dose, You may mix | blend.

Dosage A person of ordinary skill in the art can readily determine the appropriate dose of a polypeptide HP or polynucleotide sequence or host cell described herein for administration to a subject without undue experimentation. Usually, the physician will determine the actual dosage that will be most appropriate for an individual patient, including the activity of the specific bacterial strain utilized, the metabolic stability and length of action, age, It will depend on various factors including body weight, overall health, sex, diet, method and frequency of administration, excretion rate, combination of drugs, severity of the particular condition and the treatment the individual is receiving. The dosages disclosed herein are exemplary of the average case. There can, of course, be individual instances where higher or lower dosage ranges are merited, and such cases are within the scope of this invention.

Combination In one aspect, the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence are one or more It is administered in combination with another active agent. In such cases, one or more host cells comprising the polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or an expression vector comprising the polynucleotide sequence. Administration with other active agents may be continuous, simultaneous or sequential.

  For example, at least two of the polypeptide HP, polynucleotide sequences, and host cells described herein are administered to the subject.

  For example, one type of host cell according to the present invention (eg L. lactis transformed with a polynucleotide sequence encoding HP) may be transformed into another type of host cell according to the present invention (eg a polynucleotide encoding HP). Lactobacillus species transformed with the sequence) may be used in combination.

  In another example, one type of host cell according to the present invention (eg L. lactis transformed with a polynucleotide sequence encoding HP) is transformed into a Bacteroides spp. (Eg Bacteroides tetaiotaomicron), Lactococcus You may use together with another microorganism, such as genus species (for example, L. lactis), Lactobacillus species, Bifidobacterium species, and Streptococcus species (for example, Streptococcus thermophilus).

Polynucleotide Sequences The scope herein includes polynucleotide sequences that encode HP polypeptides.

  The term “nucleotide sequence” as used herein refers to oligonucleotide or polynucleotide sequences and variants, homologs, fragments and derivatives (eg, portions thereof) thereof. Nucleotide sequences may be of genomic origin, synthetic origin, or recombinant origin, which may represent the sense strand or the antisense strand. Regardless, it may be double-stranded or single-stranded.

  The term “nucleotide sequence” in this context includes genomic DNA, cDNA, synthetic DNA, and RNA. In one embodiment, the term refers to a cDNA sequence.

  In one embodiment, a nucleotide sequence when in relation to and encompassed by the original scope of the specification, refers to the native nucleotide sequence, if in the natural environment of the nucleotide sequence itself, and nucleotides It is not included when it is linked to a sequence naturally associated with the sequence (the associated sequence is also in the natural environment of the associated sequence itself). For ease of reference, this embodiment is referred to herein as a “non-native nucleotide sequence”. In this context, the term “native nucleotide sequence” is the entire nucleotide sequence that is in its own natural environment and is operative for all naturally associated promoters (the promoter is also in its own natural environment). Means the entire nucleotide sequence when linked to However, the amino acid sequences encompassed by the scope herein can be isolated and / or purified after expression of the nucleotide sequence in its native organism. However, in one embodiment, the amino acid sequence encompassed by the scope of this specification is by the nucleotide sequence in its native organism, but when the nucleotide sequence is not under the control of a naturally associated promoter in the organism, Sometimes expressed.

  Typically, the nucleotide sequences encompassed by the scope herein are prepared using recombinant DNA technology (ie, recombinant DNA). However, in alternative embodiments of the present invention, chemical methods well known in the art (Caruthers MH et al., (1980) Nuc Acids Res Symp Ser 215-23 and Horn T et al., (1980) Nuc Acids Res It is expected that nucleotide sequences can be synthesized in whole or in part using Symp Ser 225-232).

  The polynucleotides encompassed herein may be used in combination with other polynucleotide sequences. Thus, the present specification also encompasses a combination of polynucleotide sequences comprising a polynucleotide sequence encoding HP and another polynucleotide sequence that may be another polynucleotide sequence encoding HP.

Nucleotide Sequence Preparation Nucleotide sequences encoding any of the peptides herein may be identified and / or isolated and / or purified from any cell or organism that produces the peptide. Various methods for the identification and / or isolation and / or purification of nucleotide sequences are well known in the art. As an example, once the appropriate sequence has been identified and / or isolated and / or purified, DNA amplification techniques can be used to prepare more sequences.

  As a further example, genomic DNA and / or cDNA libraries can be constructed using chromosomal DNA or messenger RNA from organisms that produce peptides. If the amino acid sequence is known, labeled oligonucleotide probes can be synthesized and used to identify clones from a genomic library prepared from that organism. Alternatively, it is expected that labeled clones can be identified using labeled oligonucleotide probes containing sequences homologous to similar known genes. In the latter case, lower stringency hybridization and wash conditions are used.

  Alternatively, a clone containing the peptide of the present specification can be obtained by inserting a fragment of genomic DNA into an expression vector, such as a plasmid, transforming bacteria with the resulting genomic DNA library, and then an agar plate containing the peptide substrate. It is expected that identification can be made by allowing the identification of clones expressing the peptide by seeding the transformed bacterium into

  In yet a further alternative, the nucleotide sequence encoding the peptide is obtained from established standard methods, such as the phosphoramidite method described by Beucage SL et al., (1981) Tetrahedron Letters 22, p 1859-1869, or It can be prepared synthetically by the method described by Matthes et al., (1984) EMBO J.3, p 801-805. In the phosphoramidite method, oligonucleotides are synthesized, for example, on an automated DNA synthesizer, purified, annealed, ligated and cloned into an appropriate vector.

  Nucleotide sequences are prepared according to standard techniques (as appropriate) by synthesis, ligation of fragments of genomic or cDNA origin, mixed genomic and synthetic origin, mixed synthetic and cDNA origin, or genomic and cDNA origin. It may be of mixed origin. Each fragment that is ligated corresponds to a different part of the total nucleotide sequence. Polymerase chain reaction (PCR) using specific primers as described, for example, in US Pat. No. 4,683,202 or Saiki RK et al., (Science (1988) 239, pp 487-491). DNA sequence may be prepared by polymerase chain reaction).

Amino Acid Sequences The scope herein also includes HP polypeptides as defined herein.

  The amino acid sequence may be prepared / isolated from a suitable source, may be made synthetically, or may be prepared by use of recombinant DNA technology.

  The polypeptides encompassed herein may be used in combination with other peptides. Thus, the present description also encompasses peptide combinations comprising the polypeptide HP and another peptide that may be another HP polypeptide.

  The amino acid sequences when related to and covered by the original scope of the invention are not native peptides. In this context, the term “native peptide” means the entire peptide in its natural environment when expressed by its native nucleotide sequence.

Recombinant polypeptides In one aspect, polypeptide sequences for use in the present invention are recombinant sequences, i.e., those of a polypeptide that uses recombinant DNA technology (e.g., a host cell containing an expression vector encoding the polypeptide). Sequence prepared using (expression).

  These recombinant DNA techniques are within the ability of one skilled in the art. Such techniques are described in the literature, eg, J. Sambrook, EF Fritsch, and T. Maniatis, 1989, Molecular Cloning: A Laboratory Manual, Second Edition, Books 1-3, Cold Spring Harbor Laboratory Press.

Fusion Proteins Polypeptide sequences for use in accordance with the present invention may be produced as fusion proteins, eg, to aid in extraction and purification. Examples of fusion protein partners include glutathione-S-transferase (GST), 6xHis, GAL4 (DNA binding and / or transcriptional activation domain) and (β-galactosidase). It may be convenient to include a protein cleavage site between the fusion protein partner and the desired protein sequence to allow removal of the fusion protein sequence.

  Usually, the fusion protein will not interfere with the activity of the protein sequence.

  A gene fusion expression system in E. coli is reviewed in Curr Opin Biotechnol (1995) 6 (5): 501-6.

  In another embodiment herein, the polypeptide sequence may be ligated to a heterologous sequence to encode a fusion protein. For example, it may be useful to encode a chimeric substance that expresses a heterologous epitope recognized by a commercially available antibody to screen a peptide library for agents that can affect the activity of the substance.

Sequence identity or sequence homology The terms “polypeptide”, “polypeptide sequence”, “peptide”, “protein” and “amino acid sequence” are used synonymously herein.

  The terms “polynucleotide sequence” and “nucleotide sequence” are used synonymously herein.

  The present invention provides a degree of sequence identity or sequence homology with the amino acid sequences of the polypeptides (eg, variants, homologs and derivatives) described herein, or any nucleotide sequence encoding such polypeptides. Also included is the use of sequences having the sequence (hereinafter referred to as “homologous sequences”). As used herein, the term “homolog” means an entity having a certain homology with a subject amino acid sequence and a subject nucleotide sequence. In this specification, the term “homology” can be considered the same as “identity”.

  In this context, the homologous sequence may be at least 50, 60, 70, 75, 80, 85 or 90% identical to the subject sequence, and in some embodiments at least 95, 96, 97, 98 or 99% identical. It is taken to include amino acid or nucleotide sequences. Although homology can be considered from the viewpoint of similarity (ie, amino acid residues having similar chemical properties / functions), in the context of the present invention, it is preferable to express homology from the viewpoint of sequence identity.

  In some embodiments, a homologous sequence is taken to include an amino acid sequence or nucleotide sequence that has one or more additions, deletions and / or substitutions relative to the subject sequence.

  In some embodiments, the present invention provides a protein whose amino acid sequence is presented herein, or from this (parent) protein, one or more amino acids in the amino acid sequence of the parent protein, eg 2, 3, 4, A protein obtained by substitution, deletion or addition of 5, 6, 7, 8, 9 amino acids or more, for example 10 or more amino acids, having the activity of the parent protein About the use of.

  In some embodiments, the invention encodes a protein whose amino acid sequence is presented herein, or from this (parent) protein, one or more amino acids in the amino acid sequence of the parent protein, eg 2, 3 A protein obtained by substitution, deletion or addition of 4, 5, 6, 7, 8, 9 amino acids or more, eg, 10 or more amino acids, wherein the parent protein The use of a nucleic acid sequence (or gene) that encodes a protein having the following activity:

  In this context, a homologous sequence is at least 50, 60, 70, 75, 80, 85 or 90% identical, in some embodiments, to a nucleotide sequence (subject sequence) encoding a polypeptide described herein. It is intended to include nucleotide sequences that may be at least 95, 96, 97, 98 or 99% identical. Usually, a homolog is a sequence that encodes a domain or the like and includes a sequence that is the same as or equivalent to the target sequence. Although homology can be considered from the viewpoint of similarity (ie, amino acid residues having similar chemical properties / functions), in the context of the present invention, it is preferable to express homology from the viewpoint of sequence identity.

  Homologous amino acid sequences and / or nucleotide sequences can provide and / or encode a polypeptide that retains and / or enhances the functional activity of the polypeptide.

  In some aspects, the amino acid sequences described herein have at least 50, 60, 70, 75, 80, 85, or 90% identity with the subject sequence, in some embodiments, at least 95, 96, 97, 98 or 99% identity.

  In some aspects, the nucleotide sequence described herein has at least 50, 60, 70, 75, 80, 85, or 90% identity with the subject sequence, in some embodiments, at least 95, 96, 97, 98 or 99% identity.

  Homology comparisons can be made visually or, more usually, using readily available sequence comparison programs. These commercially available computer programs can calculate% homology between two or more sequences.

  % Homology may be calculated over consecutive sequences. That is, one sequence is aligned with another sequence and each amino acid in one sequence is directly compared one residue at a time with the corresponding amino acid in the other sequence. This is referred to as “no gap” alignment. Typically, such ungapped alignments are performed only on a relatively small number of residues.

  This is a very simple and consistent method, but for example, in the case of otherwise identical sequence pairs, subsequent amino acid residues cannot be aligned due to one insertion or deletion, and thus It cannot be taken into account that the% homology can result in a significant reduction when performing a comprehensive alignment. Therefore, most sequence comparison methods are designed to produce an optimal alignment that takes into account possible insertions and deletions without penalizing excessively the overall homology score. This is accomplished by inserting “gaps” to maximize local homology during sequence alignment.

  However, these more complex methods are such that sequence alignments using as few gaps as possible that reflect the higher association between the two compared sequences, such that the same amino acid has the same number, A “gap penalty” is assigned to each gap present in the alignment such that a score greater than that using is used. “Affine gap costs” are typically used that charge a relatively high cost for the existence of a gap and a smaller penalty for each subsequent residue in the gap. This is the most commonly used gap scoring system. Of course, a high gap penalty will produce an optimized alignment with fewer gaps. Most alignment programs can change the gap penalty. Usually, when such software is used for sequence comparison, default values are used.

  Therefore, in order to calculate the maximum homology%, it is first necessary to generate an optimal alignment that takes into account the gap penalty. A suitable computer program for performing such an alignment is Vector NTI (Invitrogen). Examples of software that can perform sequence comparison include, for example, the BLAST package (see Ausubel et al 1999 Short Protocols in Molecular Biology, 4th Ed-Chapter 18), BLAST2 (FEMS Microbiol Lett 1999 174 (2) 247-50, FEMS Microbiol Lett 1999 177 (1): 187-8 and tatiana@ncbi.nlm.nih.gov), FASTA (Altschul et al 1990 J. Mol. Biol. 403-410) and AlignX is included, but not limited thereto. At least BLAST, BLAST2, and FASTA are available for offline and online searching (see Ausubel et al 1999, pages 7-58 to 7-60).

  Although the final% homology can be measured in terms of identity, the alignment process itself is usually not based on an all-or-nothing pair comparison. Instead, a scaled similarity score matrix is generally used that assigns a score to each pair-wise comparison based on chemical similarity or evolutionary distance. An example of such a matrix that is commonly used is the BLOSUM62 matrix—the default matrix for the BLAST program suite. Vector NTI programs generally use either public default values or custom symbol comparison tables if provided (see user manual for further details). For some applications, it is preferable to use the default values for the Vector NTI package.

  Alternatively, percent homology is calculated using the Vector NTI (Invitrogen) multiple alignment mechanism based on an algorithm similar to CLUSTAL (Higgins DG & Sharp PM (1988), Gene 73 (1), 237-244). be able to.

  Once the software has produced an optimal alignment, it is possible to calculate% homology, eg% sequence identity. The software typically performs this calculation as part of the sequence comparison and generates a numerical result.

  Gap penalties should be used when determining sequence identity, in which case, for example, the following parameters can be used for pairwise alignment:

  In one embodiment, CLUSTAL may be used with a set of gap penalties and gap extensions as defined above.

  In one embodiment, the identity with respect to the nucleotide sequence is over at least 20 contiguous nucleotides, such as over at least 30 contiguous nucleotides, such as over at least 40 contiguous nucleotides, such as over at least 50 contiguous nucleotides. It is determined over at least 60 nucleotides, such as over at least 100 consecutive nucleotides, such as over at least 200 consecutive nucleotides, such as over at least 300 consecutive nucleotides.

  In one embodiment, identity with respect to the nucleotide sequence may be determined throughout the sequence.

  A sequence may have deletions, insertions or substitutions of amino acid residues that cause a static change and result in a functionally equivalent substance. If the secondary binding activity of the substance is retained, systematic amino acid substitution is performed based on similarity of residue polarity, charge, solubility, hydrophobicity, hydrophilicity and / or amphiphilicity. Also good. For example, negatively charged amino acids include aspartic acid and glutamic acid, positively charged amino acids include lysine and arginine, and amino acids having uncharged polar head groups with similar hydrophilicity values include leucine, isoleucine, valine, glycine. , Alanine, asparagine, glutamine, serine, threonine, phenylalanine and tyrosine.

  Conservative substitutions may be made, for example according to the table below. Amino acids in the same block in the second column, preferably in the same row in the third column, may be substituted for each other:

  The present invention provides for possible homologous substitutions (both substitutions and substitutions herein refer to the replacement of existing amino acid residues with alternative residues), ie, homologous substitutions, eg, between basic amino acids. Substitution, substitution between acidic amino acids, substitution between polar amino acids and the like are also included. Non-homologous substitutions, i.e., substitution of one residue from another class, or alternatively, an unnatural amino acid such as ornithine (hereinafter referred to as Z), ornithine diaminobutyrate (herein Hereinafter referred to as B), norleucine ornithine (hereinafter referred to as O), pyridylalanine, thienylalanine, naphthylalanine and phenylglycine substitution may be performed.

Also replaces there is that made by unnatural amino acids, alpha ^* and alpha-disubstituted amino acids, N- alkyl amino acids ^*, lactic acid ^*, halide derivatives of natural amino acids such as trifluorotyrosine ^*, p-Cl @ - phenylalanine ^*, p-Br-phenylalanine ^* , p-I-phenylalanine ^* , L-allyl-glycine ^* , β-alanine ^* , L-α-aminobutyric acid ^* , L-γ-aminobutyric acid ^* , L-α-aminoisobutyric acid ^* , L-ε-aminocaproic acid ^# , 7-aminoheptanoic acid ^* , L-methionine sulfone ^{# *} , L-norleucine ^* , L-norvaline ^* , p-nitro-L-phenylalanine ^* , L-hydroxyproline ^# , L-thioproline ^*, methyl derivatives of phenylalanine (Phe), such as 4-methyl -Phe , Pentamethyl -Phe ^*, L-Phe (4- ^amino) #, L-Tyr ^(methyl) *, L-Phe (4- ^isopropyl) *, L-Tic (1,2,3,4- tetrahydroisoquinoline -3 -Carboxylic acid) ^* , L-diaminopropionic acid ^# and L-Phe (4-benzyl) ^* . The notation ^* was used to indicate the hydrophobicity of the derivative for the above discussion (with respect to homologous or non-homologous substitutions), whereas # was used to indicate the hydrophilicity of the derivative. # ^* Indicates amphiphilic characteristics.

  Various amino acid sequences can be inserted between any two amino acid residues of a sequence containing an alkyl group such as a methyl, ethyl or propyl group in addition to an amino acid spacer such as a glycine or β-alanine residue. May contain a spacer group. Further variations include the presence of one or more amino acid residues in the peptoid form, which will be well understood by those skilled in the art. For the avoidance of doubt, the “peptoid form” is used to refer to variant amino acid residues in which the α carbon substituent is on the residue's nitrogen atom rather than the α carbon. Methods for preparing peptoid peptides are described in the art, for example, Simon RJ et al., PNAS (1992) 89 (20), 9367-9371 and Horwell DC, Trends Biotechnol. (1995) 13 (4), 132 -134.

  Nucleotide sequences for use in the present invention may include synthetic or modified nucleotides within the sequence. A number of different types of modifications to oligonucleotides are known in the art. These include methylphosphonate and phosphorothioate backbones and / or the addition of acridine or polylysine chains at the 3 'and / or 5' ends of the molecule. In the present invention, it is to be understood that the nucleotide sequences described herein may be modified by any method available in the art. Such modifications may be made to improve the in vivo activity or lifetime of the nucleotide sequences of the present invention.

  The invention also encompasses the use of nucleotide sequences that are complementary to the sequences presented herein or any derivative or fragment thereof. If the sequence is complementary to the fragment, the sequence can be used as a probe to identify similar coding sequences in other organisms and the like.

  Polynucleotides that are not 100% homologous to the sequences of the invention but fall within the scope of the invention can be obtained in a number of ways. Other variants of the sequences described herein may be obtained, for example, by probing DNA libraries made from a range of individuals, such as individuals from different populations. In addition, other homologues may be obtained, and such homologues and fragments thereof will generally be capable of selectively hybridizing to the sequences shown in the sequence listing herein. Such sequences may be obtained by probing cDNA libraries made from other animal species or genomic DNA libraries from other animal species, and the attached sequence listing under medium to high stringency conditions May be obtained by probing such a library with a probe comprising all or part of any one of the sequences. Similar considerations apply to obtaining species homologues and allelic variants of the polypeptide or nucleotide sequences of the invention.

  Variants and strain / species homologues may be obtained using degenerate PCR, such that the PCR targets sequences within the variants and homologues that encode conserved amino acid sequences within the sequences of the invention. The designed primer will be used. Conserved sequences can be predicted, for example, by alignment of amino acid sequences from several variants / homologs. Sequence alignment can be performed using computer software known in the art. For example, the GCG Wisconsin PileUp program is widely used.

  Primers used in degenerate PCR will contain one or more degenerate positions and have a lower stringency than that used for cloning sequences with a single sequence primer relative to a known sequence Will be used in the condition.

  Alternatively, such polynucleotides may be obtained by site-directed mutagenesis of the characterized sequence. This can be useful, for example, when static codon sequence changes are required to optimize codon preference for the particular host cell in which the polynucleotide sequence is to be expressed. Other sequence changes may be desirable in order to introduce restriction enzyme recognition sites or to alter the properties or functions of the polypeptide encoded by the polynucleotide.

  Using the polynucleotides (nucleotide sequences) of the present invention, revealing labels by conventional means using primers, eg PCR primers, primers for alternative amplification reactions, probes, eg radioactive or non-radioactive labels A probe labeled with can be produced, or the polynucleotide can be cloned into a vector. Such primers, probes and other fragments are at least 15, preferably at least 20, such as at least 25, 30 or 40 nucleotides in length, and such primers, probes and other fragments are also referred to herein. When used, it is encompassed by the term polynucleotide of the present invention.

  Polynucleotides according to the invention, such as DNA polynucleotides and probes, may be produced recombinantly, synthetically or by any means available to those skilled in the art. They may be cloned by standard techniques.

  In general, primers will be produced by synthetic means, including a step-by-step production of the desired nucleic acid sequence one nucleotide at a time. Techniques for accomplishing this production using automated techniques are readily available in the art.

  Longer polynucleotides will generally be produced using recombinant means, for example using PCR (polymerase chain reaction) cloning techniques. Primers may be designed to contain suitable restriction enzyme recognition sites so that the amplified DNA can be cloned into a suitable cloning vector.

Recombinant Polynucleotide Sequences In one aspect, a polynucleotide sequence for use in the present invention uses a recombinant polypeptide sequence, ie, a host cell comprising recombinant DNA technology (eg, an expression vector encoding the polypeptide). Polypeptide expression). Examples of recombinant polynucleotide sequences include codon optimized sequences and polynucleotide sequences encoding fusion polypeptides.

  These recombinant DNA techniques are within the ability of one skilled in the art. Such techniques are described in the literature, eg, J. Sambrook, EF Fritsch, and T. Maniatis, 1989, Molecular Cloning: A Laboratory Manual, Second Edition, Books 1-3, Cold Spring Harbor Laboratory Press.

Synthesis In one aspect, sequences for use in the present invention are synthetic sequences, ie sequences prepared by in vitro chemical or enzymatic synthesis. Such sequences include, but are not limited to, sequences made with optimal codon usage for host organisms such as the methylotrophic yeast Pichia and Hansenula.

  The invention is further illustrated by the following non-limiting examples.

[Example]
Unless otherwise indicated, the practice of the present invention will employ conventional techniques of chemistry, molecular biology, microbiology, recombinant DNA and immunology that are within the ability of one skilled in the art. Such techniques are explained in the literature. For example, J. Sambrook, EF Fritsch, and T. Maniatis, 1989, Molecular Cloning: A Laboratory Manual, Second Edition, Books 1-3, Cold Spring Harbor Laboratory Press, Ausubel, FM et al. (1995 and periodic supplements, Current Protocols in Molecular Biology, ch. 9, 13, and 16, John Wiley & Sons, New York, NY), B. Roe, J. Crabtree, and A. Kahn, 1996, DNA Isolation and Sequencing: Essential Techniques, John Wiley & Sons, JM Polak and James O'D. McGee, 1990, In Situ Hybridization: Principles and Practice, Oxford University Press; MJ Gait (Editor), 1984, Oligonucleotide Synthesis: A Practical Approach, Irl Press, DMJ Lilley and JE Dahlberg, 1992, Methods of Enzymology: DNA Structure Part A: Synthesis and Physical Analysis of DNA Methods in Enzymology, Academic Press, and EM Shevach and W. Strober, 1992 and periodic supplements, Current Protocols in Immunology, John Wiley & Sons, New See York, NY. Each of these general texts is incorporated herein by reference.

  The NF-κB luciferase reporter shows that Bacteroides tetaiotaomicron HP (BT0187, pilin related protein) greatly reduces NF-κB activity in epithelial cells in culture stimulated by flagellin-, PMA- or IL-1. Revealed by assay.

  For large-scale production, HP (referred to as HP in FIGS. 1A and 1B; FIG. 1A shows the polynucleotide sequence and FIG. 1B shows the polypeptide sequence) was transferred to E. coli or L. coli. It was expressed in lactis.

  The sequence was (i) codon optimized for expression in E. coli (referred to as Rec 1 HP in FIG. 1B); Codon optimized for expression in Lactis (referred to as Rec 2 HP in FIG. 1B).

  Isolated recombinant HP was tested in vitro and encapsulated.

  The efficacy of the encapsulated product was evaluated in a rat model of inflammatory bowel disease (dextran sulfate sodium [DSS] induced colitis).

Effect of HP on Inflammatory Bowel Disease Rat Study: Fooded-Lister rats (Rowett strain, 6 weeks old, approximately 480 g) are housed under standard quality conditions in the Bioresources of the Rowett Institute of Nutrition and Health And housed and managed. Rats were treated with sterile distilled water containing sodium dextran sulfate (MP Biomedicals UK, Cambridge; DSS; molecular weight 3600-50000) for 7 days [Days 1-5, 40 g DSS / l, and Days 6-7, 20 g DSS / l]. Half of the DSS-treated rats received HP protein daily (Days 1-7) and the remaining 6 DSS-treated rats received daily (Days 1-7) placebo. Untreated controls were given free access to sterile distilled water but were not dosed. Untreated controls received sterilized distilled water. All rats were given free access to high-quality rodent chow. Food intake, water intake and body weight were measured daily.

  On day 8, the rats were euthanized (isoflurane overdose and phlebotomy) and dissected. The total length of the colon is measured, 3-6 cm of the ascending colon from the cecum / colon junction is taken with OCT or fixed with neutral buffered formalin, 2-3 cm from the cecum / colon junction is placed in RNAlater, and the cecum / colon A 0-2 cm piece from the junction was snap frozen. Ascending colon pieces 3-6 cm from the rectum were collected by OCT or fixed with neutral buffered formalin, 2-3 cm from the rectum were placed in RNAlater, and 0-2 cm pieces from the rectum were snap frozen. Measure the small intestine, collect 5-7 cm of ileal tissue from the ileocecal junction by OCT or fix with neutral buffered formalin, collect 7-9 cm from the ileal cecal junction for microbiology, ileocecal junction 9-10 cm from was placed in RNAlater and 9-17 cm pieces from the ileocecal junction were snap frozen. The transverse colon was collected for microbiology, as well as mesenteric lymph nodes, liver and spleen. Lactose-fermenting bacteria and non-lactose-fermenting bacteria in the tissues were evaluated using MacConkey agar medium # 3.

  Fixed colon samples were embedded in technobit 8100 resin. 4 μm sections were cut and stained with hematoxylin and eosin. The entire cross-sectional area was imaged and digitized using a Zeiss Axioskop microscope connected to a QImaging camera controlled by ImageProPlus software. These were examined by two unrelated individuals in a blinded manner (Berg, DJ, Davidson, N., Kuhn, R., Muller, W., Menon, S., Holland, G., Thompson-Snipes, L ., Leach, MW, Rennick, D. Enterocolitis and colon cancer in interleukin-10-deficient mice are associated with aberrant cytokine production and CD4 + Th1-like responses (1996) Journal of Clinical Investigation, 98 (4), 1010-1020) Based on these methods, the severity of intestinal damage was categorized and the data were expressed as a percentage of the field of view for conditions from 0 [no disease] to grade 3 [serious disease].

  Rats treated with sodium dextran sulfate (DSS) and rats treated with both DSS and HP (DSS / HP) had equivalent water intake, so DSS intake was also the same between treatment groups. It was. At the same time, it was observed that food intake by DSS rats was slightly lower than that of DSS / HP treated rats and controls. DSS-treated rats also tended to lose weight, while DSS / HP and control rats maintained their weight (FIG. 2).

  Rat colon length was shortened due to DSS ingestion. This is a feature reported for DSS colitis. However, this change in the colon was prevented when the rats were also treated with HP (FIG. 3). In contrast, the length of the small intestine increased with DSS intake but did not change in rats fed DSS / HP (FIG. 3).

  The mesenteric lymph nodes, liver and spleen of rats fed DSS were asymptomatically infected with lactose (mainly E. coli) fermentable and non-lactose fermentable bacteria (FIG. 4). This was not significant with DSS / HP. The transmission of bacteria to these systemic tissues is likely the result of a loss of intestinal barrier integrity due to damage caused by DSS. HP appeared to prevent this loss of intestinal barrier integrity.

  Histological analysis of the ascending and descending colons was performed (Figures 5 and 6). (Berg, DJ, Davidson, N., Kuhn, R., Muller, W., Menon, S., Holland, G., Thompson-Snipes, L., Leach, MW, Rennick, D. Enterocolitis and colon cancer in Based on the method of interleukin-10-deficient mice are associated with aberrant cytokine production and CD4 + Th1-like responses (1996) Journal of Clinical Investigation, 98 (4), 1010-1020) Data were displayed as a percentage of the field of view for [no pathology] to grade 3 [serious pathology] pathology or as an average histopathological diagnosis score. The tissue perturbations caused by DSS were moderate and patchy, with varying degrees of damage throughout the tissue section (from little or none to severe). Overall, there was a loss of mucosal epithelial integrity, a reduction in the number of goblet cells in the epithelium, and infiltration of immune cells into the lamina propria. In contrast, general colonic disturbances caused by DSS were greatly reduced by combined treatment of rats with HP. Thus, HP was protective in the DSS-induced colitis model.

Affymetrix analysis Principal component analysis (PCA) was performed using microarray data to separate samples in three dimensions. Since the control and DSS / HP were both clustered, DSS was separated from this cluster. Therefore, this PCA showed that the DSS transcriptome profile was very different from the very similar control and DSS / HP profiles. This also showed that HP is effective in treating inflammation. It was because these animals looked like healthy animals.

  ANOVA for non-equal variance (Welch), P <0.05, asymptotic, all single conditions was performed, and Tukey HSD post-test was performed to obtain a list of differentially expressed genes. This generated a table of 377 genes with differential expression (Tables 1 and 3).

  A heat map of a subset of the 377 gene was generated (FIG. 7). This heat map showed that the DSS animals were clearly clustered away from the control and DSS / HP animals, and the control and DSS / HP animals themselves were clustered separately but not from each other. . Although the overall color patterns of the control and DSS / HP were very similar, the DSS animals showed a fold change opposite to the other two groups for the majority of this gene subset.

  Seven of these genes showed relatively high fold changes compared to controls. Four of these seven genes were up-regulated relative to the control and the other three were down-regulated (Table 1). These fold changes were generally higher for DSS animals than for DSS / HP animals. The most affected genes were upregulated relative to controls, regenerated pancreatic islet-derived 3beta (Reg3b), resistin-like gamma | resistin-like beta (Retnlg | Retnlb), sucrase-isomaltase (alpha-glucosidase) (Si ) And defensin alpha 24 (Defa24), and hydroxysteroid 11-beta dehydrogenase 2 (Hsd11b2), hydroxysteroid (17-beta) dehydrogenase 2 (Hsd17b2) and nuclear receptor 1D1 | It was hormone receptor alpha (Nr1d1 | Thra) (Table 1).

Real-time PCR
Expression of inflammation-related genes in the ascending colon was generally less in rats treated with DSS and HP than in tissues from rats treated with DSS alone (FIG. 8, Table 2). In particular, Reg3 and RELMb expression was greatly reduced as a result of HP treatment.

Summary Virtual proteins ameliorated moderate DSS-induced colitis. This protein has been linked in part to reduced expression of pro-inflammatory markers in intestinal tissue.

References
Kelly D, Campbell JI, King TP, Grant G, Jansson EA, Coutts AG, Pettersson S, Conway S. Commensal anaerobic gut bacteria attenuate inflammation by regulating nuclear-cytoplasmic shuttling of PPAR-gamma and RelA. Nat Immunol. 2004 Jan; 5 (1): 104-12
Xu J, Bjursell MK, Himrod J, Deng S, Carmichael LK, Chiang HC, Hooper LV, Gordon JI.A genomic view of the human-Bacteroides thetaiotaomicron symbiosis. Science. 2003 Mar 28; 299 (5615): 2074-6
Wendler WM, Kremmer E, Forster R, Winnacker EL.Identification of pirin, a novel highly conserved nuclear protein.J Biol Chem. 1997 Mar 28; 272 (13): 8482-9
Berg, DJ, Davidson, N., Kuhn, R., Muller, W., Menon, S., Holland, G., Thompson-Snipes, L., Leach, MW, Rennick, D. Enterocolitis and colon cancer in interleukin -10-deficient mice are associated with aberrant cytokine production and CD4 + Th1-like responses (1996) Journal of Clinical Investigation, 98 (4), 1010-1020

  All publications mentioned in the above specification are herein incorporated by reference. Various modifications and variations of the described methods and system of the invention will be apparent to those skilled in the art without departing from the scope and spirit of the invention. Although the invention has been described in connection with specific preferred embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the described modes for carrying out the invention which are obvious to those skilled in biochemistry and molecular biology or related fields are deemed to be within the scope of the following claims. .

Claims (71)

A host comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence for use in the treatment and / or prevention of a disorder of interest A cell,
The disorder is an inflammatory disorder and / or an autoimmune disorder;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence.   The disorder of claim 1, wherein the disorder affects the gastrointestinal tract, a portion of the gastrointestinal tract, liver, hepatocytes, epithelial cells, epidermal cells, neurons, kidneys, spleen, lungs, heart, pancreas, and / or pancreatic cells. A polypeptide, polynucleotide sequence, or host cell according to claim 1 for use.   Disorders include inflammatory bowel disorder (IBD), colitis, rheumatoid arthritis, psoriasis, multiple sclerosis, type I diabetes, celiac disease, atopic dermatitis, rhinitis, irritable bowel syndrome (IBS), ulcerative colitis Ileositis, Crohn's disease, functional dyspepsia, atopic disease, necrotizing enterocolitis, nonalcoholic fatty liver disease, gastrointestinal infection, lupus, nephritis / glomerulonephritis, asthma, COPD, myocarditis and combinations thereof 2. A polypeptide, polynucleotide sequence, or host cell according to claim 1 for use according to claim 1 selected from the group.   The polypeptide, polynucleotide sequence, or host cell of claim 1 for use according to claim 1, wherein the disorder is inflammatory bowel disorder (IBD). A polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or said polynucleotide sequence for use in modulating inflammation of a cell, tissue or organ of interest A host cell comprising an expression vector comprising
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence.   6. A polypeptide, polynucleotide sequence, or host cell according to claim 5 for use according to claim 5, which reduces inflammation of cells, tissues or organs.   6. The cell, tissue or organ is the digestive tract, a portion of the digestive tract, liver, hepatocyte, epithelial cell, epidermal cell, nerve cell, kidney, spleen, lung, heart, pancreas, and / or pancreatic cell. 7. A polypeptide, polynucleotide sequence or host cell according to claim 5 or 6 for use according to claim 6.   8. A polypeptide, polynucleotide sequence or host cell according to any of claims 5 to 7 for use according to any of claims 5 to 7, which reduces inflammation by epithelial cells of a tissue or organ. Including a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence for use in improving the intestinal barrier integrity of a subject A host cell,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence. Preferably for reducing the level of one or more types of lactose-fermenting bacteria in a tissue or organ of interest for use in modifying the bacterial composition in the tissue or organ to yield a beneficial microflora And / or polypeptide HP, a polynucleotide sequence encoding polypeptide HP, for use in reducing the level of one or more types of non-lactose fermenting bacteria in a tissue or organ of interest, A host cell comprising a nucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence.   11. The use according to claim 10 for use according to claim 10, wherein the tissue or organ is selected from the group consisting of mesenteric lymph nodes, liver, pancreas, spleen, kidney, heart, lung and combinations thereof. A polypeptide, polynucleotide sequence, or host cell. A polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a polynucleotide sequence for use in maintaining the length of the large and / or small intestine of a subject A host cell comprising an expression vector comprising:
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence.   13. A polypeptide, polynucleotide sequence or host cell according to claim 12 for use according to claim 12, which prevents shortening of the length of the large intestine and / or prevents an increase in length of the small intestine. A host comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence for use in reducing intestinal disturbances in a subject A cell,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence.   Reduce or prevent disruption of the integrity of the mucosal epithelium and / or reduce or prevent a reduction in the number of goblet cells in the epithelium and / or reduce or prevent infiltration of immune cells into the lamina propria 15. A polypeptide, polynucleotide sequence, or host cell according to claim 14, for use according to claim 14. A polypeptide HP for use in modulating the expression of one or more pro-inflammatory genes and / or one or more barrier integrity genes in one or more cells of a subject, A polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence. Regenerated islet-derived 3 beta gene (Reg3b), resistin-like gamma-resistin-like beta gene (Retnlg | Retnlb), sucrase-isomaltase (alpha-glucosidase) gene (Si), defensin alpha 24 gene ( Defa24), hydroxysteroid 11-beta dehydrogenase 2 gene (Hsd11b2), hydroxysteroid (17-beta) dehydrogenase 2 gene (Hsd17b2), resistin-like molecule-beta (RELMb) and nuclear receptor 1D1 thyroid hormone receptor alpha gene ( Polypeptide HP for use in regulating the expression of one or more genes selected from the group consisting of Nr1d1 | Thra) Li nucleotide sequence, a host cell comprising an expression vector comprising a host cell, or said polynucleotide sequence comprising said polynucleotide sequences,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence.   Regenerated islet-derived 3 beta gene (Reg3b), resistin-like gamma-resistin-like beta gene (Retlnlg | Retnlb), resistin-like molecule-beta (RELMb), sucrase-isomaltase (alpha-glucosidase) gene (Si), and defensin alpha 24 18. A polypeptide, polynucleotide sequence or host cell according to claim 17 for use according to claim 17, which reduces the expression of one or more genes selected from the group consisting of genes (Defa24). .   Selected from the group consisting of hydroxysteroid 11-beta dehydrogenase 2 gene (Hsd11b2), hydroxysteroid (17-beta) dehydrogenase 2 gene (Hsd17b2), and nuclear receptor 1D1 thyroid hormone receptor alpha gene (Nr1d1 | Thra) 18. A polypeptide, polynucleotide sequence, or host cell according to claim 17 for use according to claim 17, which increases the expression of one or more genes. A polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, for use in reducing activation of a pro-inflammatory pathway in one or more cells of a subject, or A host cell comprising an expression vector comprising a polynucleotide sequence,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence. A polypeptide HP for use in reducing the activity and / or expression of NF-κβ in one or more cells of interest, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, Or a host cell comprising an expression vector comprising said polynucleotide sequence,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence. A polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or an expression vector comprising the polynucleotide sequence for use in improving the health of the subject's gastrointestinal tract A host cell,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
A host cell comprising said polypeptide HP, a polynucleotide sequence encoding said polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence.   23. A polypeptide, polynucleotide sequence, or host cell according to any of claims 1 to 22 for use according to any of claims 1 to 22, which is encapsulated.   24. A polypeptide according to any of claims 1 to 23 for use according to any of claims 1 to 23, which is a recombinant polypeptide.   24. A polynucleotide sequence according to any of claims 1 to 23 for use according to any of claims 1 to 23, wherein the expression vector comprises a polynucleotide sequence. A polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence, and a pharmaceutically acceptable excipient, carrier or A pharmaceutical composition comprising a diluent,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said pharmaceutical composition.   27. The pharmaceutical composition of claim 26, wherein the polypeptide, polynucleotide sequence, or host cell is encapsulated.   28. The pharmaceutical composition according to claim 26 or 27, wherein the polypeptide is a recombinant polypeptide.   28. A pharmaceutical composition according to claim 26 or 27, wherein the expression vector comprises a polynucleotide sequence. A polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence and a nutritionally acceptable excipient, carrier or A nutritional supplement containing a diluent,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
The nutritional supplement.   31. The nutritional supplement of claim 30, wherein the polypeptide, polynucleotide sequence, or host cell is encapsulated.   The nutritional supplement according to claim 30 or 31, wherein the polypeptide is a recombinant polypeptide.   32. A nutritional supplement according to claim 30 or 31, wherein the expression vector comprises a polynucleotide sequence. Veterinary feed, food, dietary supplement or food additive comprising polypeptide HP, polynucleotide sequence encoding polypeptide HP, host cell comprising said polynucleotide sequence, or host cell comprising an expression vector comprising said polynucleotide sequence A thing,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
The livestock feed, food, nutritional supplement, or food additive.   35. Livestock feed, food, dietary supplement or food additive according to claim 34 which is encapsulated.   36. Livestock feed, food, dietary supplement or food additive according to claim 34 or 35, wherein the polypeptide is a recombinant polypeptide.   36. Livestock feed, food, dietary supplement or food additive according to claim 34 or 35, wherein the expression vector comprises a polynucleotide sequence. 30. A method for producing a pharmaceutical composition according to any of claims 26 to 29, wherein the polypeptide, polynucleotide sequence or host cell is mixed with a pharmaceutically acceptable excipient, carrier or diluent. Including steps;
The polypeptide, polynucleotide sequence, or host cell may be encapsulated in the manner;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method for producing the nutritional supplement according to any one of claims 30 to 33, comprising:
Mixing the polypeptide, polynucleotide sequence, or host cell with a nutritionally acceptable excipient, carrier or diluent;
The polypeptide or polynucleotide may be encapsulated by the method;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method for producing a livestock feed, food, dietary supplement or food additive according to any of claims 34-37,
Mixing the polypeptide, polynucleotide sequence, or host cell with its livestock feed, food, dietary supplement, food additive, or ingredient;
The polypeptide or polynucleotide may be encapsulated by the method;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method for treating and / or preventing a disorder in a subject comprising:
Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence;
Said polypeptide, polynucleotide sequence, or host cell treats and / or prevents said subject's disorder;
The disorder is an inflammatory disorder and / or an autoimmune disorder;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method of modulating inflammation of a tissue or organ of interest comprising:
Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence;
The polypeptide, polynucleotide sequence, or host cell modulates inflammation of the tissue or organ of interest;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method for improving a subject's intestinal barrier integrity, comprising:
Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence;
The polypeptide, polynucleotide sequence, or host cell improves intestinal barrier integrity of the subject;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method of reducing the level of one or more types of lactose-fermenting bacteria in a subject tissue or organ and / or reducing the level of one or more types of lactose-fermenting bacteria in a target tissue or organ There,
Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence;
The polypeptide, polynucleotide sequence, or host cell reduces the level of one or more types of lactose-fermenting bacteria in the tissue or organ of interest and / or 1 or 2 in the tissue or organ of interest. Reduce the level of these types of non-lactose fermenting bacteria;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method of maintaining the length of a subject's large intestine and / or small intestine, comprising:
Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence;
The polypeptide, polynucleotide sequence, or host cell maintains the length of the large and / or small intestine of the subject;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method for reducing disturbances in a subject's intestines,
Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence;
The polypeptide, polynucleotide sequence, or host cell reduces intestinal disturbances in the subject;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method of modulating the expression of one or more pro-inflammatory genes in one or more cells of a subject comprising:
Administering to the subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, or a host cell comprising the polynucleotide sequence, or a host cell comprising an expression vector comprising the polynucleotide sequence;
The polypeptide, polynucleotide sequence, or host cell modulates the expression of one or more pro-inflammatory genes in the one or more cells of the subject;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method of modulating the expression of one or more genes in one or more cells of a subject comprising:
Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence;
The polypeptide, polynucleotide sequence, or host cell regulates the expression of one or more genes in the one or more cells of the subject;
The one or more genes include a regenerated islet-derived 3beta gene (Reg3b), a resistin-like gamma-resistin-like beta gene (Retnlg | Retnlb), a sucrase-isomaltase (alpha-glucosidase) gene (Si), a defensin alpha 24 genes (Defa24), hydroxysteroid 11-beta dehydrogenase 2 gene (Hsd11b2), hydroxysteroid (17-beta) dehydrogenase 2 gene (Hsd17b2), resistin-like molecule-beta (RELMb), and nuclear receptor 1D1 thyroid hormone receptor Selected from the group consisting of a body alpha gene (Nr1d1 | Thra);
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Said method has at least 75% identity with a variant, homolog, fragment or derivative;   A method for reducing activation of a pro-inflammatory pathway in one or more cells of a subject, comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or said polynucleotide Administering to said subject a host cell comprising an expression vector comprising the sequence, wherein said polypeptide or polynucleotide sequence or host cell reduces activation of a pro-inflammatory pathway in said one or more cells of said subject. The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or SEQ ID NO: 6 or a variant, homologue, fragment or derivative thereof Encodes a polypeptide having at least 75% identity and / or the polynucleotide sequence is at least 75% identical to SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof The method having a sex. A method for reducing NF-κB activity and / or expression in one or more cells of a subject comprising:
Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence;
The polypeptide or polynucleotide sequence or host cell reduces the activity and / or expression of NF-κβ in the cell or cells of the subject;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. A method for improving the health of the subject's digestive tract,
Administering to said subject a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence;
The polypeptide or polynucleotide sequence, or host cell, improves the digestive tract health of the subject;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said method. Polypeptide HP, polynucleotide sequence encoding polypeptide HP, host cell comprising the polynucleotide sequence, or expression vector comprising the polynucleotide sequence for the manufacture of a medicament for the treatment and / or prevention of a disorder of interest Using a host cell comprising
The disorder is an inflammatory disorder and / or an autoimmune disorder;
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said use. Polypeptide HP, polynucleotide sequence encoding polypeptide HP, host cell comprising the polynucleotide sequence, or expression vector comprising the polynucleotide sequence for producing a drug that modulates inflammation of a target tissue or organ Using a host cell comprising
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said use. Including a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or an expression vector comprising the polynucleotide sequence for producing a drug that improves the intestinal barrier integrity of a subject Use of a host cell,
Said polypeptide has at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof;
The polynucleotide sequence encodes a polypeptide having at least 75% identity with SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologues, fragments or derivatives thereof, and / or the polynucleotide sequence comprises: Has at least 75% identity with SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or variants, homologues, fragments or derivatives thereof;
Said use. A polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell containing the polynucleotide sequence, or a polynucleotide sequence for producing a drug that maintains the length of the large and / or small intestine of a subject Use of a host cell comprising an expression vector comprising:
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said use. A drug that reduces the level of one or more types of lactose-fermenting bacteria in a subject tissue or organ and / or reduces the level of one or more types of non-lactose-fermenting bacteria in a target tissue or organ Using a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, comprising:
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said use. A host comprising a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, or an expression vector comprising the polynucleotide sequence for producing a drug that reduces intestinal disturbances in the subject Cell use,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said use. Polypeptide HP, poly for producing a drug that modulates the expression of one or more pro-inflammatory genes and / or one or more barrier integrity genes in one or more cells of a subject Use of a polynucleotide sequence encoding peptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said use. Regenerated islet-derived 3 beta gene (Reg3b), resistin-like gamma-resistin-like beta gene (Retnlg | Retnlb), sucrase-isomaltase (alpha-glucosidase) gene (Si), defensin alpha 24 gene ( Defa24), hydroxysteroid 11-beta dehydrogenase 2 gene (Hsd11b2), hydroxysteroid (17-beta) dehydrogenase 2 gene (Hsd17b2), resistin-like molecule-beta (RELMb) and nuclear receptor 1D1 thyroid hormone receptor alpha gene ( Nr1d1 | Thra), which encodes polypeptide HP for producing a drug that regulates the expression of one or more genes selected from the group consisting of Polynucleotide sequence, to the use of a host cell comprising an expression vector comprising a host cell, or said polynucleotide sequence comprising said polynucleotide sequences,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said use. Polypeptide HP, a polynucleotide sequence encoding polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising said polynucleotide sequence, for producing a drug that reduces activation of a pro-inflammatory pathway in one or more cells of a subject Use of a host cell comprising an expression vector comprising a polynucleotide sequence comprising
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said use. A polypeptide HP for producing a drug that reduces the activity and / or expression of NF-κβ in one or more cells of interest, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising the polynucleotide sequence, Or the use of a host cell comprising an expression vector comprising said polynucleotide sequence,
The polypeptide has at least 75% identity to SEQ ID NO: 2, SEQ ID NO: 4 or SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof; and the polynucleotide sequence is SEQ ID NO: 2, SEQ ID NO: 4 Or encodes a polypeptide having at least 75% identity with SEQ ID NO: 6 or variants, homologs, fragments or derivatives thereof, and / or wherein said polynucleotide sequence is SEQ ID NO: 1, SEQ ID NO: 3 or SEQ ID NO: 5 or Have at least 75% identity with a variant, homolog, fragment or derivative;
Said use.   A polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell containing the polynucleotide sequence, or an expression vector containing the polynucleotide sequence for producing a drug that improves the health of the subject's gastrointestinal tract Use of host cells.   Including a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or an expression vector comprising said polynucleotide sequence for use substantially as described herein Host cell.   A pharmaceutical composition substantially as described herein.   A nutritional supplement substantially as described herein.   Livestock feed, food, dietary supplement or food additive substantially as described herein.   A method of manufacturing a pharmaceutical composition substantially as described herein.   A method of manufacturing a nutritional composition substantially as described herein.   A method of producing livestock feed, food, dietary supplement or food additive substantially as described herein.   Use of a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, substantially as described herein Method.   Use of a polypeptide HP, a polynucleotide sequence encoding the polypeptide HP, a host cell comprising said polynucleotide sequence, or a host cell comprising an expression vector comprising said polynucleotide sequence, substantially as described herein .